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BIOCHEMISTRY

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TEXTBOOK OF

BIOCHEMISTRY

For Dental Students

Second Edition

DM Vasudevan

MBBS MD FAMS FRC (Path) Distinguished Professor of Biochemistry

Amrita Institute of Medical Sciences, Kochi, Kerala, India

Principal (Retd), College of Medicine, Amrita Institute of Medical Sciences, Kochi, Kerala, India Formerly, Dean, Sikkim Manipal Institute of Medical Sciences, Gangtok, Sikkim, India

Sreekumari S MBBS MD

Professor, Department of Biochemistry

Sree Gokulam Medical College and Research Foundation Thiruvananthapuram, Kerala, India

Kannan Vaidyanathan MBBS MD

Associate Professor and Head

Department of Biochemistry Metabolic Disorders Laboratory

Amrita Institute of Medical Sciences, Kochi, Kerala, India

JAYPEE BROTHERS MEDICAL PUBLISHERS (P) LTD

New Delhi • Panama City • London ®

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Headquarter

Jaypee Brothers Medical Publishers (P) Ltd 4838/24, Ansari Road, Daryaganj

New Delhi 110 002, India Phone: +91-11-43574357 Fax: +91-11-43574314

Email: jaypee@jaypeebrothers.com

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Website: www.jaypeebrothers.com Website: www.jaypeedigital.com

© 2011, Jaypee Brothers Medical Publishers

All rights reserved. No part of this book and DVD ROM may be reproduced in any form or by any means without the prior permission of the publisher.

Inquiries for bulk sales may be solicited at: jaypee@jaypeebrothers.com

This book has been published in good faith that the contents provided by the author(s) contained herein are original, and is intended for educational purposes only. While every effort is made to ensure a accuracy of information, the publisher and the author(s) specifically disclaim any damage, liability, or loss incurred, directly or indirectly, from the use or application of any of the contents of this work. If not specifically stated, all figures and tables are courtesy of the authors(s). Where appropriate, the readers should consult with a specialist or contact the manufacturer of the drug or device.

Publisher: Jitendar P Vij

Publishing Director: Tarun Duneja Editor: Richa Saxena

Cover Design: Seema Dogra, Sumit Kumar

Textbook of Biochemistry for Dental Students

First Edition: 2007 Second Edition: 2011 ISBN 978-93-5025-488-2 Printed in India

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Dedicated to

At the lotus feet of the Holy Mother,

Sri Mata Amritanandamaji Devi

"Today's world needs people who express goodness in their words and deeds. If such noble role models set the example for their fellow beings, the darkness prevailing in today's society will be dispelled, and the light of peace and non-violence will once again illumine this earth. Let us work together towards this goal."

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We are glad to present the second edition of the Textbook of Biochemistry for Dental Students. Many medical colleges and universities in India have accepted the first edition of the book as one of the standard textbooks. With humility, we may state that the medical community of India has warmly received the previous edition of this book. In retrospect, it gives immense satisfaction to note that the book served the students and faculty for the past few years.

At this time, a revision of the textbook has become absolutely necessary because the Dental Council of India have revised the syllabus for biochemistry. This book is prepared according the syllabi (modified in 2007) prepared by the Dental Council of India. We have also consulted the syllabi of dental courses of various universities.

In this edition, we have deleted some extra details shown in the first edition. We have also added one chapter on Endocrinology, as it is now introduced by the Dental Council of India. In this edition, important points are made into bold letters, so that students can grasp the subject easily. Moreover, the facts are given in pointwise (with numbers), so as to aid easy memorization. The essentials of biochemistry are given in ordinary font, the "must know points" will be available from these paragraphs. A small group of students who aspire for more knowledge, a bit more advanced topics are given in small prints. Students just before the examination can skip the tiny letters, but these areas will also become important later in their pursuit of knowledge. Those who are interested to get more facts may consult our Textbook of

Biochemistry for Medical Students, now in 6th edition. A question bank, given in the end of the book

contains essay questions and short note questions, which are compiled from the question papers of various universities during the last decade. These questions will be ideal for students for last-minute preparation for examinations.

The first author is in the process of retirement from active service, and would like to reduce the burden in due course. A successful textbook is something like a growing institution; individuals may come and go, but the institution will march ahead. Therefore, we felt the need to induce younger blood into the editorial board. Thus, a third author has been added in this edition, so that the torch can be handed over smoothly at an appropriate time later on.

The first author desires more interaction with faculty and students who are using this textbook. All are welcome to communicate at his email address <dmvasudevan@yahoo.co.in>. The help and assistance rendered by our students in preparing this book are enormous. Web pictures, without copyright protection were used in some figures. The remarkable success of the book was due to the active support of the publishers. This is to record our appreciation for the cooperation extended by Shri Jitendar P Vij, Chairman and Managing Director, Jaypee Brothers Medical Publishers Pvt Ltd, New Delhi and his associates.

We hope that this edition will be friendlier to the students and be more attractive to the teachers. Now this is in your hands to judge.

“End of all knowledge must be building up of character.”—Gandhiji

DM Vasudevan Sreekumari S Kannan Vaidyanathan

Preface to the Second Edition

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The medical community of India has warmly received the "Textbook of Biochemistry for Medical Students", edited by us. It was originally published in 1995, and now running the 4th edition. We were getting persistent requests from teachers of dental courses that a concise version of the Textbook is necessary for catering their needs. In order to satisfy this continued demand, we are now presenting this Textbook of

Biochemistry for Dental Students.

This book is prepared after consulting the syllabi of Dental courses of various universities. As there are slight variations on the emphasis given by different universities, we have tried to incorporate all the facts prescribed in all those curricula. So, some students may find some chapters unnecessary for their immediate purpose. But a general textbook of this nature has to satisfy the needs of all students.

The important points are made into bold letters, so that students can grasp the subject easily. Moreover, the facts are given in point-wise (with numbers), so as to aid easy memorization. There are two sets of undergraduate students; majority wants only a pass. For them, the essentials of Biochemistry are given in ordinary font. A small group of students aspire for distinction; for them a bit more advanced knowledge is given in small prints. Students just before the examination can skip the tiny letters, but these areas will also become important later in their pursuit of knowledge. Those who are interested to get more facts may consult our Textbook of Biochemistry for Medical Students.

A question bank, given in the end of the book contains essay questions, short notes and viva voce type questions. These questions are compiled from the question papers of various universities during the last decade. These questions will be ideal for students for last-minute preparation for examinations.

The first author desires more interaction with faculty and students who are using this textbook. All are welcome to communicate at his e-mail address <dmvasudevan@aims.amrita.edu>.

We extend our thanks for the enormous help rendered by Dr Ananth N Rao and Dr R Krishnaprasad in preparation of this book. We also thank Shri Jitendar P Vij, Chairman and Managing Director, Jaypee Brothers Medical Publishers Pvt Ltd, New Delhi and his associates for their active support in making this publication possible.

A textbook will be matured only by successive revisions. We hope that this first edition will be friendly to the students and be attractive to the teachers. Now this is in your hands to judge.

“End of all knowledge must be building up of character.”—Gandhiji

DM Vasudevan Sreekumari S

Preface to the First Edition

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1. Subcellular Organelles and Cell Membranes ... 1 Introduction 1, Biomolecules 1, Subcellular Organelles 1, Nucleus 2, Endoplasmic

Reticulum (ER) 2, Golgi Apparatus 3, Lysosomes 3, Mitochondria 3, Plasma Membrane 3, Cytoskeleton 4, Transport Mechanisms 4, Simple Diffusion 4, Facilitated Diffusion 4, Ion Channels 4, Ligand Gated Channels 5, Amelogenin 5, Voltage Gated Channels 5, Active Transport 5, Uniport, Symport and Antiport 5

2. Amino Acids and Proteins ... 7 Classification of Amino Acids 7, Based on Structure 7, Special Groups in Amino Acids 9,

Classification Based on Side Chain 9, Classification Based on Metabolic Fate 9, Classification Based on Nutritional Requirement 9, Properties of Amino Acids 10, Isoelectric Point 10, Optical Activity 10, Reactions Due to Carboxyl Group 11, Reactions Due to Amino Group 11, Reactions Due to Side Chains 11, Special Functions of Amino Acids 12, Color Reactions of Amino Acids and Proteins 12, Peptide Bond 12, Structure of Proteins (Organization of Proteins) 13, Primary Structure; Sequence 13, Secondary Structure of Proteins 13, Tertiary Structure 14, Quaternary Structure 14, Structure-function Relationship 15, Physical Properties of Proteins 15, Isoelectric pH (PI) of Proteins 15, Precipitation Reactions 15, Denaturation of Proteins 16, Heat Coagulation 16, Classification of Proteins 16, Classification Based on Functions 16, Classification Based on Solubility 16, Classification Based on Nutritional Value 17

3. Enzymology ... 19 Classification of Enzymes 19, IUBMB System of Nomenclature of Enzymes 19,

Coenzymes 19, First Group of Coenzymes 20, Nicotinamide Adenine Dinucleotide (NAD+) 20, Second Group of Coenzymes 20, Adenosine Triphosphate (ATP) 20, Salient Features of Coenzymes 21, Metallo-enzymes 21, Mode of Action of Enzymes 21, Lowering of Activation Energy 21, Michaelis-menten Theory 21, Fischer's Template Theory 22, Koshland's Induced Fit Theory 22, Active Site or Active Center 22, Thermodynamics 22, Exergonic or Exothermic Reaction 22, Isothermic Reaction 22, Endergonic or Endothermic Reaction 22, Enzyme Kinetics 23, Factors Influencing Enzyme Activity 23, Enzyme Concentration 23, Substrate Concentration 23, Effect of Concentration of Products 24, Effect of Temperature 24, Effect of pH 24, Enzyme Activation 24, Covalent Modification 25, Enzyme Inhibition 25, Competitive Inhibition 25, Noncompetitive Inhibition 26, Allosteric Regulation 26, Key Enzymes 27, Induction 27, Repression 27, Isoenzymes 27, Clinical Enzymology 28, Lactate Dehydrogenase (LDH) 28, Creatine Kinase (CK) 28, CK and Muscle Diseases 29, Alanine Amino Transferase (ALT) 29, Alkaline Phosphatase (ALP) 29

4. Carbohydrates–I: Chemistry, Digestion and Absorption of Carbohydrates... 31 Functions of Carbohydrates 31, Nomenclature 31, Stereoisomers 31, D and L Isomerism

of Glucose 32, Optical Activity 32, Diastereoisomers of Glucose 32, Glucose, Mannose and Galactose 32, Epimerism of Aldoses 33, Anomerism of Sugars 33, Three Representations of Glucose Structure 33, Fructose is a Ketohexose 34, Reactions of Monosaccharides 34, Enediol Formation 34, Benedict's Reaction 34, Osazone Formation 34, Reduction to Form Alcohols 35, Oxidation of Sugars 35,

Contents

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Furfural Derivatives 35, Glycosides 35, Formation of Esters 36, Amino Sugars 36, Deoxy Sugars 36, Pentoses 36, Disaccharides 36, Sucrose 36, Lactose 37, Maltose 37, Isomaltose 37, Polysaccharides 37, Starch 38, Glycogen 38, Cellulose 38, Inulin 38, Chitin 38, Heteroglycans 39, Agar 39, Mucopolysaccharides 39, Hyaluronic Acid 39, Heparin 39, Keratan Sulphate 39, Glycoproteins and Mucoproteins 39, Digestion of Carbohydrates 39, Lactose Intolerance 39, Absorption of Carbohydrates 40, Co-transport from Lumen to Intestinal Cell 40, Release of Glucose Into Blood 40, GluT2 in Other Tissues 40, GluT4 in Muscle and Adipose Tissue 40

5. Carbohydrates–II: Major Metabolic Pathways of Glucose ... 42 Glucose Metabolism 42, Importance of Glucose 42, Glycolysis 42, Embden-Meyerhof

Pathway 42, Importance of the Pathway 42, Step 0: Glucose Entry Into Cells 42, Step 1 of Glycolysis 42, Step 2 of Glycolysis 43, Step 3 of Glycolysis 43, Step 4 of Glycolysis 43, Step 5 of Glycolysis 44, Step 6 of Glycolysis 44, Step 7 of Glycolysis 44, Step 8 of Glycolysis 44, Step 9 of Glycolysis 44, Step 10 of Glycolysis 44, Steps 5 and 10 are Coupled 45, Energy Yield from Glycolysis 45, Cori's Cycle or Lactic Acid Cycle 45, Regulation of Glycolysis 46, Metabolic Fate of Pyruvate 47, Pyruvate Dehydrogenase Complex 47, Pyruvate as a Junction Point 47, Gluconeogenesis 47, Pyruvate Carboxylase Reaction 47, Phosphoenolpyruvate Carboxykinase 47, Reversal of Glycolysis 48, Fructose-1,6-bisphosphatase 48, Glucose-6-phosphatase 48, Energy Requirement for Gluconeogenesis 48, Substrates for Gluconeogenesis 48, Regulation of Gluconeogenesis 49, Hormonal Regulation of Gluconeogenesis 50, Physiological Significance 50, Glycogen Metabolism 50, Functions of Glycogen 50, Degradation of Glycogen (Glycogenolysis) 50, Glycogen Synthesis (Glycogenesis) 51, Regulation of Glycogen Metabolism 51, Glycogen Storage Diseases 52

6. Carbohydrates–III: Regulation of Blood Sugar, Diabetes Mellitus ... 54 Regulation of Blood Glucose 54, Factors Maintaining Blood Sugar 54, Postprandial

Regulation 54, Regulation in Fasting State 54, Normal Plasma Glucose Level 55, Sugar in Urine 55, Determination of Glucose in Body Fluids 56, Effect of Food on Glucose Level 56, Oral Glucose Tolerance Test (OGTT) 56, Normal Values and Interpretations 57, Diagnostic Criteria for Diabetes Mellitus 57, Causes for Abnormal GTT Curve 57, Reducing Substances in Urine 57, Identification of Reducing Sugars 58, Diabetes Mellitus 58, Type 1 Diabetes Mellitus 58, Type 2 Diabetes Mellitus 58, Metabolic Derangements in Diabetes 59, Clinical Presentations in Diabetes Mellitus 59, Laboratory Investigations in Diabetes 60, Management of Diabetes Mellitus 60

7. Carbohydrates–IV: Minor Metabolic Pathways of Carbohydrates ... 61 Hexose Monophosphate (HMP) Shunt Pathway 61, Overview of the Shunt Pathway 61,

Oxidative Phase 61, Nonoxidative Phase 62, Regulation of HMP Shunt Pathway 62, Significance of the HMP Shunt Pathway 62, Glucuronic Acid Pathway 62, Importance of the Glucuronic Acid Pathway 62, Vitamin C in Lower Animals 63, Essential Pentosuria 63, Fructose Metabolism 63, Hereditary Fructose Intolerance (HFI) 63, Fructosuria 64, Galactose Metabolism 64, Galactose Catabolism 64, Galactosemia 64, Metabolism of Alcohol 65, Alcohol Dehydrogenase (ADH) 65, Aldehyde Dehydrogenase 65, Biochemical Alterations in Alcoholism 65, Mucopolysaccharidoses 65

8. Biochemistry of Teeth, Saliva and Dental Caries ... 67 Functions of Saliva 67, Composition of Saliva 67, Inorganic Components 68, Organic

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Trace Elements 70, Organic Components 70, Proteins of Dentin 71, Dental Caries 71, Microbiological Organisms Cause Dental Caries 71, Fluoride 73, Fluoride is Useful to Prevent Caries 74, Fluorosis is More Dangerous than Caries 74, Incidence of Fluorosis 75, Prevention of Fluorosis 75

9. Lipids–I: Chemistry ... 76 Functions of Lipids 76, Clinical Applications 76, Classification of Lipids 76, Simple

Lipids 76,Compound Lipids 76, Derived Lipids 77, Lipids Complexed to Other Compounds 77, Fatty Acids 77, Classification of Fatty Acids 77, Depending on Total No. of Carbon Atoms 77, Depending on Length of Hydrocarbon Chain 77, Depending on Nature of Hydrocarbon Chain 77, Saturated Fatty Acids 77, Unsaturated Fatty Acids 78, Clinical Significance of PUFA 78, Properties of Fatty Acids 78, Neutral Fats 78, Phospholipids 79, Phosphatidates 79, Amphipathic Nature 80, Biomembranes 80, Liposomes 80, Nonphosphorylated Lipids 80, Lipoproteins 80, Lipid Storage Diseases or Sphingolipidoses 81, Niemann Pick's Disease 81, Gaucher's Disease 81, Tay-Sachs Disease 81, Essential Fatty Acids 81, Eicosanoids 81, Prostaglandins (PGS) 81, Biosynthesis of Prostaglandins 81, Regulation of Synthesis 81, Biological Actions 82

10. Lipids–II: Metabolism of Fatty Acids ... 83 Digestion of Lipids 83, Digestion in Stomach 83, Digestion in Intestines 83, Digestion

of Triglycerides 83, Absorption of Lipids 84, Absorption of Long Chain Fatty Acids 84, Beta Oxidation of Fatty Acids 85, Preparative Step for Beta Oxidation 85, Beta Oxidation Steps 86, Oxidation of Odd Chain Fatty Acids 87, Propionate is Gluconeogenic 87, Inborn Errors of Propionate Metabolism 87, De Novo Synthesis of Fatty Acids 88, Transport of Acetyl CoA to Cytoplasm 88, Fatty Acid Synthase (FAS) Complex 88, Coenzymes of Fatty Acid Synthesis 89, Regulation of Fatty Acid Synthesis 89, Synthesis of Triglycerides (TAG) 90, Obesity 90, Fatty Liver and Lipotropic Factors 90, Causes of Fatty Liver 91, Lipotropic Factors 91, Metabolism of Ketone Bodies 91, Ketogenesis 91, Ketolysis 93, Ketosis 93, Causes for Ketosis 93, Regulation of Ketogenesis 93, Diabetes, Starvation, Ketosis, Cholesterol 94, Consequences of Ketosis 94, Diagnosis of Ketosis 94, Differential Diagnosis 94, Management of Ketoacidosis 94

11. Lipids–III: Cholesterol, Lipoproteins and Cardiovascular Diseases ... 96 Significance and Functions of Cholesterol 96, Structure of Cholesterol 96, Biosynthesis

of Cholesterol 96, Regulation of Cholesterol Synthesis 97, Excretion of Cholesterol 98, Plasma Lipids 98, Carriage of Cholesterol in Blood 98, Classification of Lipoproteins 98, General Characteristics of Lipoproteins 99, Apolipoproteins 99, Chylomicrons 99, Very Low Density Lipoproteins 100, Low Density Lipoproteins (LDL) 100, Metabolism of LDL and LDL Receptors 100, Function of LDL 100, LDL and Clinical Applications 101, High Density Lipoprotein (HDL) 101, Metabolism of HDL 101, Function of HDL 102, Clinical Significance of HDL 102, Free Fatty Acid (FFA) 102, Hyperlipidemias 102, Type IIA (Primary Familial Hypercholesterolemia) 103, Atherosclerosis 103, Atherosclerosis and LDL 104, Serum Cholesterol is Increased in 104, Risk Factors for Atherosclerosis 104, Serum Cholesterol Level 104, LDL-cholesterol Level 104, HDL-cholesterol Level 104, Apoprotein Levels and Ratios 105, LP(a) 105, Cigarette Smoke 105, Hypertension 105, Diabetes Mellitus 105, Serum Triglyceride 105, Obesity and Sedentary Life Style 105, Prevention of Atherosclerosis 105, Reduce Dietary Cholesterol 105, Vegetable Oils and PUFA 105, Moderation in Fat Intake 105, Plenty of Green Leafy Vegetables 105, Avoid Sucrose and Cigarette 105, Exercise 105, Hypolipidemic Drugs 105

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12. Amino Acid Metabolism ... 107 Digestion of Proteins 107, Gastric Digestion of Proteins 108, Pancreatic Digestion of

Proteins 108, Intestinal Digestion of Proteins 108, Absorption of Amino Acids 108, Clinical Applications 108, General Metabolism of Amino Acids 109, Formation of Ammonia 109, Transamination 109, Trans-deamination 109, Disposal/Detoxification of Ammonia 109, First Line of Defence (Trapping of Ammonia) 109, Final Disposal 110, Urea Cycle 110, Step 1: Formation of Carbamoyl Phosphate 110, Step 2: Formation of Citrulline 110, Step 3: Formation of Argininosuccinate 110, Step 4: Formation of Arginine 111, Step 5: Formation of Urea 111, Regulation of the Urea Cycle 111, Disorders of Urea Cycle 111, Urea Level in Blood and Urine 111, Glycine (GLY) (G) 111, Glycine Cleavage System 111, Special Metabolic Functions of Glycine 111, Creatine and Creatine Phosphate 112, Glycine as a Conjugating Agent 112, Methionine (Met) (M) 112, Functions of Cysteine 113, Metabolic Functions of Glutathione 113, Homocystinuria 113, Phenylalanine (PHE) (F) and Tyrosine (TYR) (T) 113, Phenylalanine to Tyrosine 113, Catabolism of Tyrosine and Phenylalanine 113, Important Specialized Products from Tyrosine 114, Phenylketonuria (PKU) 115, Laboratory Diagnosis 115, Treatment of Phenylketonuria 115, Alkaptonuria 115, Albinism 116, Tryptophan (TRP) (W) 116, Nicotinic Acid Pathway of Tryptophan 116, Serotonin 116, Melatonin 117, Histidine (His) (H) 117, Histamine 117, Biogenic Amines 117, One-carbon Metabolism 118

13. Plasma Proteins ... 122 Electrophoresis 122, Normal Values and Interpretations 122, Abnormal Patterns in

Clinical Diseases 122, Albumin 123, Functions of Albumin 123, Albumin-globulin Ratio 124, Transport Proteins 124, Acute Phase Proteins 124, C-reactive Protein (CRP) 124, Ceruloplasmin 124, Wilson's Disease 124, Structure of Immunoglobulins 125, Heavy and Light Chains 125, Variable and Constant Regions 125, Different Classes of Immunoglobulins 125, Paraproteinemias 126, Multiple Myeloma (Plasmacytoma) 126, Bence Jones Proteinuria 126, Hypergammaglobulinemia 126

14. Citric Acid Cycle, Biological Oxidation and Electron Transport Chain ... 128 Functions of the Citric Acid Cycle 128, Reactions of the Cycle 128, Significance of

TCA Cycle 131, Regulation of the Citric Acid Cycle 132, Inhibitors of TCA Cycle 132, Stages of Oxidation of Food Stuffs 133, First Stage 133, Second Stage 133, Third Stage 133, Redox Potentials 133, Substrate Level Phosphorylation 133, Biological Oxidation 133, Electron Transport Chain 133, Energetics of Oxidation Phosphorylation 133, NAD+ Linked Dehydrogenases 133, Fad-linked Dehydrogenases 134, High Energy

Compounds 134, Adenosine Triphosphate (ATP) 134, Organization of ET Chain 134, Sites of ATP Synthesis 135, Chemi-osmotic Theory 135, ATP Synthase (5th Complex) 136, Energetics of ATP Synthesis 136, Inhibitors of ATP Synthesis 136

15. Heme and Hemoglobin ... 137 Structure of Hemoglobin 137, Structure of Heme 137,Biosynthesis of Heme 137,

Regulation of Heme Synthesis 139, Disorders of Heme Synthesis 139, Acute Intermittent Porphyria (AIP) 139, Congenital Erythropoietic Porphyria 139, Diagnosis and Treatment of Porphyrias 139, Acquired Porphyrias 139, Catabolism of Heme 139, Generation of Bilirubin 139, Transport to Liver 140, Conjugation in Liver 140, Excretion of Bilirubin to Bile 140, Fate of Conjugated Bilirubin in Intestine 140, Enterohepatic Circulation 140, Final Excretion 140, Plasma Bilirubin 140, van den Bergh Test for Bilirubin 141, Hyperbilirubinemias 141, Congenital Hyperbilirubinemias 141, Acquired

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Hyperbilirubinemias 141, Hemolytic Jaundice 141, Hepatocellular Jaundice 142, Obstructive Jaundice 142, Structure of Hemoglobin 142, Transport of Oxygen by Hemoglobin 142, Oxygen Dissociation Curve (ODC) 143, Transport of Carbon Dioxide 144, Fetal Hemoglobin (HBF) 145, Hemoglobin Derivatives 145, Carboxy-hemoglobin (Carbon Monoxy-HB) (Co-HB) 145, hemoglobin (HB) 145, Met-hemoglobinemias 145, Hemoglobin (Globin Chain) Variants Hemoglobinopathies 146, Hemoglobin S (HbS) 146, Hemoglobin E 146, Thalassemias 146, Beta Thalassemia 147, Thalassemia Syndromes 147, Myoglobin (MB) 147, Anemias 147, Anemias Due to Impaired Production of RBCs 147, Hemolytic Anemias Due to Intracorpuscular Defect 147, Hemolytic Anemias Due to Extracorpuscular Causes 147, Hemorrhage 147

16. Fat Soluble Vitamins (A, D, E and K) ... 149 Vitamin A 149, Chemistry 149, Absorption of Vitamin A 149, Transport from Liver to

Tissues 149, Uptake by Tissues 150, Biochemical Role of Vitamin A 150, Dark Adaptation Mechanism 150, Rods are for Vision in Dim Light 150, Cones are for Color Vision 151, Biochemical Functions of Vitamin A 151, Deficiency Manifestations of Vitamin A 151, Causes for Vitamin A Deficiency 151, Dietary Sources of Vitamin A 151, Daily Requirements of Vitamin A 152, Hypervitaminosis A or Toxicity 152, Vitamin D (Cholecalciferol) 152, Formation of Vitamin D 152, Activation of Vitamin D 152, Biochemical Effects of Vitamin D 152, Deficiency of Vitamin D 153, Requirements of Vitamin D 153, Sources of Vitamin D 153, Hypervitaminosis D 153, Vitamin E 154, Chemical Nature 154, Biochemical Role of Vitamin E 154, Inter-relationship with Selenium 154, Deficiency Manifestations of Vitamin E 154, Sources of Vitamin E 154, Requirement 154, Vitamin K 154, Chemistry of Vitamin K 154, Biochemical Role of Vitamin K 154, Causes for Deficiency of Vitamin K 154, Clinical Manifestations of Deficiency 154, Daily Requirements of Vitamin K 155, Sources of Vitamin K 155 17. Water Soluble Vitamins (Thiamine, Riboflavin, Niacin, Pyridoxine,

Pantothenic Acid, Biotin, Folic Acid, Vitamin B12 and Ascorbic Acid) ... 156

B Complex Group of Vitamins 156, Thiamine (Vitamin B1) 156, Sources 156,

Physiological Role of Thiamine 156, Deficiency Manifestations of Thiamine 156, Recommended Daily Allowance of Thiamine 157, Riboflavin (Vitamin B2) 157, Structure

of Riboflavin 157, Coenzyme Activity of Riboflavin 157, Riboflavin Deficiency 157, Dietary Sources of Riboflavin 157, Daily Requirement 157, Niacin 157, Chemistry of Niacin 157, One Hydrogen Atom and One Electron 158, NAD+ Dependent Enzymes 158,

NADPH Reactions 158, Niacin Deficiency 158, Niacin is Synthesized from Tryptophan 158, Causes for Niacin Deficiency 159, Dietary Sources of Niacin 159, Recommended Daily Allowance (RDA) 159, Vitamin B6 159, Coenzyme form 159, Functions of Pyridoxal

Phosphate 159, Deficiency Manifestations of Pyridoxine 160, Dietary Sources 160, Requirement of B6 160, Pantothenic Acid 160, Structure 160, Coenzyme Activity of

Pantothenic Acid 160, Deficiency of Pantothenic Acid 161, Sources of Pantothenic Acid 161, Requirement of Pantothenic Acid 161, Biotin 161, Biotin Requiring Co2

Fixation Reactions 161, Biotin-independent Carboxylation Reactions 161, Biotin Antagonists 161, Requirement of Biotin 161, Sources of Biotin 161, Folic Acid 161, Chemistry of Folic Acid 161, Coenzyme Functions of Folic Acid 161, Causes for Folate Deficiency 162, Deficiency Manifestations 162, Sources of Folic Acid 162, Recommended Daily Allowance (RDA) 163, Folate Antagonists 163, Vitamin B12 163,

Chemistry 163, Absorption of Vitamin B12 163, Functional Role of B12 163, Causes of

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Dietary Sources 164, Ascorbic Acid (Vitamin C) 164, Chemistry of Vitamin C 164, Biosynthesis of Ascorbic Acid in Animals 164, Excretion of Ascorbic Acid 164, Biochemical Functions of Vitamin C 165, Deficiency Manifestations of Vitamin C 165, Dietary Sources of Vitamin C 165, Requirement of Vitamin C 165, Therapeutic Use of Vitamin C 165

18. Mineral Metabolism ... 167 Calcium (Ca++) 167, Sources of Calcium 167, Daily Requirement of Calcium 167,

Absorption of Calcium 167, Functions of Calcium 167, Calcium in Blood 168, Factors Regulating Blood Calcium Level 168, Hypercalcemia 169, Hypocalcemia and Tetany 169, Osteoporosis 171, Osteopetrosis 171, Paget’s Disease 171, Markers of Bone Diseases 171, Osteocalcin 171, Phosphorus 171, Functions of Phosphate Ions 171, Requirement and Source 171, Serum Level of Phosphorus 171, Sodium (Na+) 172, Edema 172,

Causes of Hyponatremia 172, Potassium (K+) 172, Requirement 172, Sources 172,

Normal Level 172, Hypokalemia 172, Hyperkalemia 172, Chloride (Cl) 173,

Hyperchloremia is seen in 173, Causes for Hypochloremia 173, Iron (Fe) 173, Distribution of Iron 173, Requirement of Iron (ICMR, 1990) 173, Sources of Iron 173, Factors Influencing Absorption of Iron 173, Mucosal Block Theory 173, Regulation of Absorption by 4 mechanisms 174, Iron Transport in Blood 174, Storage of Iron 174, Iron is Conserved 174, Excretion of Iron 175, Iron Deficiency Anemia 175, Causes for Iron Deficiency 175, Manifestations of Iron Deficiency Anemia 175, Treatment of Iron Deficiency 175, Iron Toxicity 175, Copper (Cu) 176, Functions of Copper 176, Abnormal Metabolism of Copper 176, Zinc (Zn) 176, Zinc Deficiency Manifestations 176, Requirement of Zinc 176, Iodine 176, Heavy Metal Poisons 176, Lead Poisoning 176

19. Energy Metabolism and Nutrition ... 178 Calorific Value 178, Respiratory Quotient (RQ) 178, Energy Requirements of a Normal

Person 178, Basal Metabolic Rate (BMR) 178, or Resting Metabolic Rate (RMR) 178, Factors Affecting BMR 178, Normal Value for BMR 179, Specific Dynamic Action (SDA) 179, Physical Activity 179, Requirements of Dietary Nutrients 179, Proximate Principles 180, Importance of Carbohydrates 180, Dietary Carbohydrates 180, Sucrose 180, Dietary Fiber 180, Nutritional Importance of Lipids 180, Cholesterol and Heart Diseases 180, Recommended Daily Intake of Lipids 181, Importance of Proteins 181, Nitrogen Balance 181, Factors Affecting Nitrogen Balance 181, Maintenance of Nitrogen Balance 182, Nutritional Values (Nutritional Indices) 182, Limiting Amino Acids 182, Supplementation 182, Protein-energy Malnutrition 182, Biochemical Alterations 183, Treatment of Protein Energy Malnutrition 183, Obesity 183, Diseases Related with Obesity 183, Prescription of Diet 183, First Step: Calorie Requirement 183, Second Step: Proximate Principles 183, Third Step: General Composition of Food 184, Mutual Supplementation of Cereals and Pulses 184, Fourth Step: Determine the Items of Food 184

20. Detoxification and Free Radicals ... 186 Enzyme Systems 186, Phases of Detoxification Processes 186, Phase One Reactions 186,

Oxidative Reactions 186, Reductive Reactions 187, Hydrolysis 187, Phase Two Reactions: Conjugations 187, Glucuronic Acid 187, Sulfate Conjugation 188, Cysteine and Glutathione 188, Acetylation 188, Conjugation with Glycine 188, Methylation Reactions 188, Free Radicals 188, Generation of Free Radicals 188, Free Radical Scavenger Enzyme Systems 189, Damage Produced by Reactive Oxygen Species 189, Clinical Significance 189, Antioxidants 189

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21. Acid-Base, pH, Electrolyte and Water Balance ... 190 Acids and Bases 190, Definition 190, Weak and Strong Acids 190, Acidity of a Solution

and pH 190, Henderson-Hasselbalch Equation 190, Buffers 190, Definition 190, Composition of a Buffer 191, Factors Affecting pH of a Buffer 191, Factors Affecting Buffer Capacity 191, How do Buffers Act? 191, Effective Range of a Buffer 191, Acid-base Balance 191, Normal pH 191, Acidosis 191, Alkalosis 191, Volatile and Fixed Acids 191, Mechanisms of Regulation of pH 191, Buffers of the Body Fluids 191, Bicarbonate Buffer System 191, Alkali Reserve 192, Phosphate Buffer System 192, Buffers Act Quickly, but Not Permanently 192, Respiratory Regulation of pH 192, The Second Line of Defense 192, Renal Regulation of pH 192, Excretion of H+; Generation of Bicarbonate 193, Reabsorption of Bicarbonate 193, Excretion of H+ as Titratable Acid 193, Excretion of Ammonium Ions 193, Relationship of pH with K+ Ion Balance 194, Classification of Acid-base Disturbances 194, Metabolic Acidosis 194, Metabolic Alkalosis 194, Respiratory Acidosis 194, Respiratory Alkalosis 195, Normal Serum Electrolyte Values 195, Electrolyte and Water Balance 195, Intake and Output of Water 195, Osmolality of Extracellular Fluid 195, Regulation of Sodium and Water Balance 196, Renin-angiotensin System 196, Autoregulation 196, Disturbances in Fluid and Electrolyte Balance 196

22. Muscle and Supportive Tissue Proteins ... 198 Collagen 198, Structure of Collagen 198, Synthesis of Collagen 198, Hydroxylation of

Proline and Lysine 198, Triple Stranded Helix 198, Quarter Staggered Arrangement 198, Cross Links in Collagen Fibers 199, Functions of Collagen 199, Abnormalities in Collagen 199, Elastin 199, Keratins 199, Muscle Proteins 199, Myosin 200, Actin 200

23. Nucleotides: Chemistry and Metabolism ... 202 Composition of Nucleotides 202, Bases Present in the Nucleic Acids 202, Nucleosides 202,

Nucleotides 203, Nucleoside Triphosphates 204, Biosynthesis of Purine Nucleotides 205, Step 0 (Preparatory Step), PRPP Synthesis 205, De Novo Synthesis of Purine Nucleotides 205, Formation of AMP 205, Conversion of IMP to GMP 205, Salvage Pathway 205, Regulation of Purine Synthesis 205, Analogs as Purine Synthesis Inhibitors 206, Degradation of Purine Nucleotides 206, Uric Acid 206, Disorders of Purine Metabolism 206, Gout 206, Primary Gout 206, Secondary Hyperuricemia 207, Clinical Findings of Gout 207, Treatment Policies in Gout 207, Lesch-Nyhan Syndrome 207, De Novo Synthesis of Pyrimidine 207, Regulation of Pyrimidine Synthesis 207, Disorders of Pyrimidine Metabolism 208, Orotic Aciduria 208, Synthesis of Deoxythymine Nucleotides 208

24. DNA: Structure and Replication ... 209 Structure of DNA 209, Polarity of DNA Molecule 209, Watson-Crick Model of DNA

Structure 210, Higher Organization of DNA 210, Nucleosomes 210, Further Condensation of DNA 211, Chromosomes 211, DNA is a Very Big Molecule 211, Inactivation of DNA during Differentiation 211, Introns, Exons, Cistrons 211, Replication of DNA 211, Steps of Replication 211, DNA Polymerase (DNAP) 212, Initiation of DNA Replication 212, RNA Primer is Required for DNA Synthesis 212, Elongation of DNA Strand 213, Discontinuous Synthesis 213, Lagging Strand and Okazaki Pieces 213, Inhibitors of DNA Replication 213

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25. Transcription and Translation ... 215 Ribonucleic Acid 215, Central Dogma of Molecular Biology 215, Replication,

Transcription and Translation 215, Template and Coding Strands 216, Messenger RNA or mRNA 216, Promoters 216, Transcription Process 216, Inhibitors of RNA Synthesis 217, Protein Biosynthesis 217, Transfer RNA (tRNA) or (sRNA) 218, Ribosomal RNA (rRNA) 218, Genetic Code 218, Translation Process 219, Inhibitors of Protein Synthesis 221

26. Control of Gene Expression ... 222 Mutations 222, Classification of Mutations 222, Effects of Mutations 222, Mutagens

and Mutagenesis 223, Cell Cycle 223, Regulation of Gene Expression 224, Operon Concept of Gene Regulation 224, The Lac Operon 224, Transcription is Normally Repressed 224, Derepression of Lac Operon 224, Clinical Applications 225, Regulation of Genes by Repression 225, Hormone Response Elements (HRE) 226

27. Recombinant DNA Technology and Gene Therapy ... 227 Recombinant DNA Technology 227, Applications of Recombinant Technology 227,

Restriction E nd on ucle ases (RE ) 2 27 , Ve ctors 22 8, P ro ced ure o f DNA Recombination 228, Human Genome Project (HGP) 229, Gene Therapy 229, Summary of the Procedure 229, How the Genes are Introduced? 230, The Vectors 230, Accomplishments 230, Stem Cells 230

28. AIDS and Cancer ... 231 AIDS and HIV 231, The Indian Scene 231, Transmission 231, Natural Course of the

Disease 231, Pathology of HIV and AIDS 232, Cancer 233, Etiology of Cancer 233, Mutagens 233, Aflatoxins 233, Cigarette 233, Antimutagens 233, Oncogenic Viruses 234, Oncogenes 234, Oncogenes are Normal Constituents of Cells 234, Many Factors Activate Oncogenes 234, Anti-oncogenes or Onco-suppressor Genes 235, Oncofetal Antigens 235, Tumor Markers 235, Clinically Important Tumor Markers 236

29. Methods of Separation and Assay of Biological Compounds... 237 Electrophoresis 237, Factors Affecting Electrophoresis 237, Electrophoresis

Apparatus 237, Cellulose Acetate Membrane 237, Agar or Agarose 237, Visualization of Protein Bands 237, Immunoelectrophoresis 238, Chromatography 238, Partition Chromatography 238, Paper Chromatography 238, Thin Layer Chromatography (TLC) 238, Visualization of Chromatography 238, Importance of Rf Value 239, Hybridization and Blot Techniques 239, Probes 239, DNA-DNA Hybridization 239, Southern Blot Technique 239, Northern Blotting for Identifying RNA 240, Western Blot Analysis for Proteins 240, Polymerase Chain Reaction (PCR) 240, Clinical Applications of PCR 240, Radioimmunoassay (RIA) 240, Advantage of RIA 241, Disadvantages of RIA 241, ELISA Test 241, Antibody Detection by ELISA 241, Antigen Detection by ELISA Method 242, Colorimeter 242, Spectrophotometer 243

30. Liver, Kidney and Gastric Function Tests ... 244 Markers of Hepatic Dysfunction 244, Measurement of Bilirubin (Test of Excretory

Function of Liver) 244, Serum Bilirubin 244, Urinary Bilirubin 245, Urinary Urobilinogen 245, Causes of Jaundice 245, Tests Based on Synthetic Function of Liver 246, Serum Albumin Level 246, Prothrombin Time (PT) 246, Alpha Feto Protein (AFP) 246, Tests Based on Serum Enzymes (Liver Enzyme Panel) 246, Enzyme Indicating Hepatocellular Damage 246, Markers of Obstructive Liver Disease 246,

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Renal Function Tests 247, Glomerular Function 247, Glomerular Filtration Rate (GFR) 247, Functions of the Tubules 247, Renal Threshold 247, Reabsorption of Water 248, Nonprotein Nitrogen (NPN) 248, Markers of GFR 249, Clearance Tests 249, Definition 249, Creatinine Clearance Test 249, Normal Serum Creatinine Level 250, Creatinine Clearance Test 250, Reference Values for Creatinine Clearance 250, Interpretation of Creatinine Clearance 250, Urea Clearance Test 250, Blood Urea Level 250, Markers of Glomerular Permeability 250, Tests for Tubular Function 251, Specific Gravity of Urine 251, Concentration Test 251, Dilution Tests 251, Urinary Acidification 251, Gastric Function 251, Mechanism of HCl Secretion 251, Regulation of Acid Secretion 251, Assessment of Gastric Function 251, Pentagastrin Stimulation Test 252, Interpretations of Gastric Juice Analysis 252, Estimation of Free Acidity and Total Acidity 252, Augmented Histamine Test 252

31. Endocrinology ... 253 Hormones Acting through Cyclic AMP (cAMP) 253, Signal Transduction through

G-protein 253, G-protein Activates Adenyl Cyclase 253, Subunit Activation of G-protein 254, Inactivation 254, Second Messenger Activates Protein Kinases 254, Protein Kinase Phosphorylates the Enzymes 254, There are Many G-proteins 255, There are Many Protein Kinases 255, Phosphatases 255, Action of Hormones Acting through Calcium 255, Hormones Acting through PIP2 Cascade 255, Role of Cyclic Gmp 255, Hormones with Intracellular Receptors 255, Adrenal Cortical Hormones 256, Synthesis of Steroid Hormones 256, Secretion of Adrenal Hormones 256, Biological Effects of Glucocorticoids 256, Assessment of Glucocorticoid Secretion 256, Assessment of Adrenal Androgen Secretion 257, Sex Hormones 257, Ovarian Hormones 257, Regulation of Ovarian Hormones 258, Testicular Hormones 258, Thyroid Hormones 258, Synthesis of Thyroxine 258, Metabolic Effects of Thyroid Hormones 259, Assessment of Thyroid Function 259, Hyperthyroidism 260, Hypothyroidism 261, Insulin 261, Structure of Insulin 261, Biosynthesis of Insulin 261, Factors Increasing Insulin Secretion 261, Physiological Actions of Insulin or Mechanisms of Action of Insulin or Metabolic Effects of Insulin 262, Hyperglycemic Hormones 263, Glucagon 263, Physiological Actions of Glucagon 263, Other Important Hormones 264

Appendices ... 265 Abbreviations Used in this Book 265

Normal Values 267

Recommended Daily Allowance (RDA) of Essential Nutrients 268 Composition of Nutrients in Selected Common Food Materials 269

Essay Questions and Short Notes ... 271

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CHAPTER AT A GLANCE

The reader will be able to answer questions on the following topics: 1. Nucleus 2. Endoplasmic reticulum 3. Golgi apparatus 4. Lysosomes 5. Mitochondria 6. Plasma membrane 7. Transport mechanisms 8. Simple and facilitated diffusion 9. Ion channels

10. Active transport

11. Uniport, symport and antiport

INTRODUCTION

Biochemistry is the language of biology. The tools for research in all the branches of medical science are mainly biochemical in nature. The study of biochemistry is essential to understand basic functions of the body. How the food that we eat is digested, absorbed, and used to make ingredients of the body? How does the body derive energy for the normal day to day work? How are the various metabolic processes interrelated? What is the function of genes? Answer for such basic questions can only be derived by a systematic study of biochemistry.

Modern day medical practice is highly dependent on the laboratory analysis of body fluids, especially the blood. The disease manifestations are reflected in the composition of blood and other body fluids. The study of biochemistry is necessary to give the scientific basis for disease and is useful for intelligent treatment of patients.

The practice of medicine is both an art and a science. The word "doctor" is derived from the Latin root, "docere", which means "to teach". The word chemistry is derived from the Greek word "chemi" (the black land), the ancient name of Egypt, when science was called the "black art". Indian medical science, even from ancient times, had identified the metabolic and genetic basis of diseases. Charaka, the great master of Indian Medicine, in his treatise (circa 400 BC) observed that madhumeha

(diabetes mellitus) is produced by the alterations in the metabolisms of carbohydrates and fats; the statement still holds good.

The term "Biochemistry" was coined by Neuberg in 1903 from Greek words, bios (= life) and chymos (= juice). Some of the important milestones in the development of science of biochemistry are given in Table 1.1. Biochemistry is the most rapidly developing subject in medicine. Thanks to the advent of DNA-recombination technology, genes can now be transferred from one person to another, so that many of the genetically determined diseases are now amenable to gene therapy.

BIOMOLECULES

More than 99% of the human body is composed of 6 elements, i.e. oxygen, carbon, hydrogen, nitrogen, calcium and phosphorus. Human body is composed of about 60% water, 15% proteins, 15% lipids, 2% carbohydrates and 8% minerals.

In living organisms, biomolecules are ordered into a hierarchy of increasing molecular complexity. These biomolecules are covalently linked to each other to form macromolecules of the cell, e.g. glucose to glycogen, amino acids to proteins, etc. Major complex biomolecules are Proteins, Polysaccharides, Lipids and Nucleic acids. The macromolecules associate with each other to form supramolecular systems, e.g. ribosomes, lipoproteins. Finally at the highest level of organization in the hierarchy of cell structure, various supramolecular complexes are further assembled into cell organelle. In prokaryotes (bacteria; Greek word "pro" = before; karyon = nucleus), these macromolecules are seen in a homogeneous matrix; but in eukaryotic cells (higher organisms; Greek word "eu" = true), the cytoplasm contains various subcellular organelles. Comparison of prokaryotes and eukaryotes are shown in Table 1.2.

SUBCELLULAR ORGANELLES

When the cell membrane is disrupted, the organized particles inside the cell are externalized. These are called subcellular organelles. They are described below.

Subcellular Organelles

and Cell Membranes

1

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1. Nucleus

a. It is the most prominent organelle of the cell. All cells in the body contain nucleus, except mature RBCs in circulation. In some cells, nucleus occupies most of the available space, e.g. small lymphocytes and spermatozoa (Fig. 1.1).

b. Nucleus is surrounded by two membranes: the inner one is called perinuclear membrane with numerous pores. The outer membrane is conti-nuous with membrane of endoplasmic reticulum. c. Nucleus contains the DNA, the chemical basis of genes which governs all the functions of the cell . DNA molecules are complexed with proteins

to form chromosomes. DNA replication and RNA synthesis (transcription) are taking place inside the nucleus.

d. In some cells, a portion of the nucleus may be seen as lighter shaded area; this is called nucleolus. This is the area for RNA processing and ribosome synthesis. The nucleolus is very prominent in cells actively synthesizing proteins. 2. Endoplasmic Reticulum (ER)

a. It is a network of interconnecting membranes enclosing channels or cisternae, that are continuous from perinuclear envelope to outer plasma membrane. Under electron microscope, the reticular arrangements will have railway track appearance (Fig. 1.1).

b. This will be very prominent in cells actively synthesizing proteins, e.g. immunoglobulin secreting plasma cells. The proteins, glycoproteins Table 1.1: Important milestones in the history

of biochemistry

Scientists Year Landmark discoveries

Rouell 1773 Isolated urea from urine Lavoisier 1785 Oxidation of food stuffs

Wohler 1828 Synthesis of urea

Louis Pasteur 1860 Fermentation process Edward Buchner 1897 Extracted the enzymes Fiske & Subbarao 1929 Isolated ATP from muscle

Lohmann 1932 Creatine phosphate

Hans Krebs 1937 Citric acid cycle Avery & Macleod 1944 DNA is genetic material Watson & Crick 1953 Structure of DNA Nirenberg & Matthai 1961 Genetic code in mRNA

Holley 1963 Sequenced gene for tRNA

Khorana 1965 Synthesized the gene Paul Berg 1972 Recombinant DNA technology Kary Mullis 1985 Polymerase chain reaction

2003 Human gene mapping

Fig. 1.1: A typical cell

Table 1.2: Comparison between prokaryotic cells and eukaryotic cells

Prokaryotic Eukaryotic cell cell

Size Small Large; 1000 to

10,000 times

Cell wall Rigid Membrane of

lipid bilayer

Nucleus Not defined Well-defined

Organelles including Nil Several mitochondria

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and lipoproteins are synthesized in the ER. Moreover, enzyme present in ER, cytochrome P-450 will detoxify various drugs.

3. Golgi Apparatus

a. It may be considered as the converging area of ER. While moving through ER, carbohydrate groups are successively added to the nascent proteins (Fig. 1.1).

b. These glycoproteins finally reach the Golgi bodies. The carbohydrate chains are further added in the Golgi prior to secretion.

c. Main function of Golgi apparatus is protein sorting, packaging and secretion.

4. Lysosomes

a. Solid wastes of a township are usually decom-posed in incinerators. Inside a cell, such a process is taking place within the lysosomes. They are bags of enzymes (Fig. 1.1).

b. Lysosomes contain enzymes that hydrolyse polysaccharides, lipids, proteins, and nucleic acids.

c. Endocytic vesicles and phagosomes are fused with lysosome (primary) to form the secondary lysosome or digestive vacuole. Foreign particles are progressively digested inside these vacuoles. 5. Mitochondria

a. They are spherical, oval or rod-like bodies, about 0.5–1 micrometers in diameter and up to 7 micrometers in length (Fig. 1.1). Erythrocytes do not contain mitochondria. The tail of spermatozoa is fully packed with mitochondria.

b. Mitochondria are the powerhouse of the cell, where energy released from oxidation of food stuffs is trapped as chemical energy in the form of ATP (see Chapter 14).

c. Mitochondria have two membranes. The inner membrane convolutes into folds or cristae. The mitochondrial membrane contains the enzymes of electron transport chain. The fluid matrix contains the enzymes of citric acid cycle. d. Mitochondria also contain specific DNA which

encodes information for synthesis for certain mitochondrial proteins. The division of mito-chondria is under the command of mitomito-chondrial DNA.

e. Antibiotics inhibiting bacterial protein synthesis do not affect cellular processes, but do inhibit

mitochondrial protein biosynthesis. Functions of organelles are shown in Tables 1.3 and 1.4. 6. Plasma Membrane

a. The plasma membrane separates the cell from the external environment. The membranes also separate different parts of the cell from one another, so that cellular activities are compartmentalized. It has highly selective permeability properties so that the entry and exit of compounds are regulated. The membrane is very active metabolically.

b. Membranes are mainly made up of lipids, proteins and small amount of carbohydrates. The contents of these compounds vary according to the nature of the membrane. The carbohydrates are present as glycoproteins and glycolipids. Phospholipids are the most common lipids present and they are amphipathic in nature. Cell membranes contain cholesterol also.

c. Fluid mosaic model: Membranes are made up of lipid bilayer. The phospholipids are arranged in bilayers with the polar head groups oriented towards the extracellular side and the cytoplasmic side with a hydrophobic core (Fig. 1.2). Each Table 1.3: Metabolic functions of subcellular

organelles

Nucleus DNA replication, transcription

Endoplasmic Biosynthesis of proteins, glycoproteins, reticulum lipoproteins, drug metabolism.

Golgi body Maturation of synthesized proteins Lysosome Degradation of proteins, carbohydrates,

lipids and nucleotides

Mitochondria Electron transport chain, ATP generation, TCA cycle, beta oxidation of fatty acids, ketone body production

Cytosol Protein synthesis, glycolysis, glycogen metabolism, transaminations, fatty acid synthesis.

Table 1.4: Comparison of cell with a factory

Plasma : Fence with gates; gates open membrane when message is received Nucleus : Manager’s office

Endoplasmic : Conveyer belt of production units reticulum

Golgi apparatus : Packing units Lysosomes : Incinerators

Vacuoles : Lorries carrying finished products Mitochondria : Power generating units

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leaflet is 25 Å thick. The total thickness is about 50 to 80 Å.

d. The lipid bilayer shows free lateral movement of its components, hence the membrane is said to be fluid in nature. However, the components do not freely move from inner to outer layer, or outer to inner layer (flip-flop movement is restricted). Fluidity enables the membrane to perform endocytosis and exocytosis.

e. Membrane proteins: The peripheral proteins exist on the surfaces of the bilayer. They are attached by ionic and polar bonds to polar heads of the lipids.

f. The integral membrane proteins are deeply embedded in the bilayer. Some of the integral membrane proteins span the whole bilayer and they are called transmembrane proteins. They can serve as receptors (for hormones, growth factors, neurotransmitters), tissue specific antigens, ion channels, membrane based enzymes, etc. 7. Cytoskeleton

Human body is supported by the skeletal system; similarly the structure of a cell is maintained by the cytoskeleton present underneath the plasma membrane. The cytoskeleton is made up of a network of microtubules and microfilaments, which contain the proteins spectrin and ankyrin. Tubules consist of polymers of tubulin.

TRANSPORT MECHANISMS

The permeability of substances across cell membranes is dependent on their solubility in lipids

and not on their molecular size. Water soluble compounds are generally impermeable and require carrier mediated transport. Transport mechanisms are classified into:

a. Passive transport i. Simple diffusion ii. Facilitated diffusion b. Active transport

c. Ion channels allow passage of molecules in accordance with the concentration gradient d. Pumps can drive molecules against the

gradient using energy. 1. Simple Diffusion

Solutes and gases enter into the cells passively. They are driven by the concentration gradient. The rate of entry is proportional to the solubility of that solute in the hydrophobic core of the membrane. Simple diffusion occurs from higher to lower concen-tration. This does not require any energy. But, it is a very slow process.

2. Facilitated Diffusion

This is a carrier mediated process (Fig.1.3C). Important features of facilitated diffusion are:

i. Structurally similar solutes can competitively inhibit the entry of the solutes.

ii. This mechanism does not require energy but the rate of transport is more rapid than diffusion process. It is dependent on concentration gradient. iii. Hormones regulate the number of carrier

molecules.

iv. Example of facilitated transport of glucose across membrane is by glucose transporters. Transport of glucose into adipose tissue and skeletal muscle is insulin dependent.

3. Ion Channels

i. Membranes have special devices called ion channels for quick transport of electrolytes such as Ca++, K+, Na+ and Cl--. These are selective ion

conductive pores. Ion channels are specialized protein molecules that span the membranes. ii. Cation conductive channels generally remain

closed but in response to stimulus, they open allowing rapid flux of ions down the gradient. This may be compared to opening of the gate of a cinema house, when people rush to enter. Hence this regulation is named as "gated". (Fig. 1.3B).

iii. Based on the nature of stimuli that trigger the opening of the gate, they are classified into "voltage gated" or "ligand gated" ion channels. Fig. 1.2: The fluid mosaic model of membrane

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Voltage gated channels are opened by membrane depolarization. Ligand gated channels are opened by binding of effectors. a. Ligand Gated Channels

Acetylcholine receptor is an example for ligand gated ion

channel. Acetylcholine released from the presynaptic region binds with the receptors on the postsynaptic region, which triggers opening of the channel and influx of Na+. This generates an action potential in the postsynaptic nerve. The channel opens only for a millisecond, because the acetylcholine is rapidly degraded by acetylcholinesterase.

b. Amelogenin

It is a protein present in enamel of teeth has hydrophobic residues on the outside. It acts as a calcium channel, which helps in mineralization (see Chapter 8).

c. Voltage Gated Channels

The channel is usually closed in the ground state. The membrane potential change (voltage difference) switches the ion channel to open. Voltage gated sodium channels and vo ltag e g ate d p o tassiu m c h an n els ar e th e c o mmo n examples.

4. Active Transport

a. The salient features of active transport are: i. This form of transport requires energy. About

40% of the total energy expenditure in a cell is used for the active transport system. (Fig.1.3A).

ii. It requires specialized integral proteins called transporters. The transporters are susceptible to inhibition by specific organic or inorganic compounds.

b. Cell has low intracellular sodium; but concentration of potassium inside the cell is very high. This is maintained by the sodium—potassium activated ATPase, generally called as sodium pump. The ATPase is an integral protein of the membrane. The hydrolysis of one molecule of ATP can result in expulsion of 3 Na+ ions and influx of 2 K+ ions.

The ion transport and ATP hydrolysis are tightly coupled.

Clinical applications: Cardiotonic drug digoxin inhibits the sodium—potassium pump. This leads to an increase in Na+ level inside the cell and

extrusion of Ca+ from the myocardial cell. This would

enhance the contractility of the cardiac muscle and so improve the function of the heart. Calcium and potassium channel blockers are widely used in therapy.

c. Calcium pump: The ATP dependent calcium pump also functions to regulate muscle contraction. A specialized membrane system called sarcoplasmic reticulum is found in skeletal muscles which regulates the Ca++ concentration

around muscle fibers. In resting muscle the concentration of Ca++ around muscle fibers is

low. But stimulation by a nerve impulse results in a sudden release of large amounts of Ca++. This

would trigger muscle contraction. The function of calcium pump is to remove cytosolic calcium and maintain low cytosolic concentration, so that muscle can receive the next signal. For each ATP hydrolyzed, 2Ca++ ions are transported.

5. Uniport, Symport and Antiport

Transport systems are classified as uniport, symport and antiport systems:

Fig. 1.3: Transport mechanisms

Table 1.5. Summary: Transport mechanisms

Carrier Energy Examples required

Simple diffusion Nil Nil Water

Facilitated Yes Nil Glucose to RBCs

diffusion

Primary Yes Directly Sodium pump

active

Secondary Yes Indirect Glucose to intestine active

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a. Uniport system (Fig.1.3D) carries single solute across the membrane, e.g. glucose transporter in most of the cells. Calcium pump is another example.

b. If the transfer of one molecule depends on simultaneous or sequential transfer of another molecule, it is called

co-transport system. The active co-transport may be coupled

with energy indirectly. Here, movement of the substance against a concentration gradient is coupled with movement of a second substance down the concentration gradient; the second molecule being already concentrated within the cell by an energy requiring process.

c. The cotransport system may either be a symport or an antiport. In symport (Fig.1.3E), the transporter carries two solutes in the same direction across the membrane, e.g. sodium dependent glucose transport.

d. The antiport system (Fig.1.3F) carries two solutes or ions in opposite direction, e.g. sodium pump or chloride-bicarbonate exchange in RBC (see Chapter 15). Features of transport modalities are summarized in Table 1.5. Exocytosis

Secretory vesicles or vacuoles move towards and fuse with the plasma membrane. This movement is created by cytoplasmic contractile elements; the microtubule system. The inner membrane of the vesicle fuses with outer plasma membrane, while cytoplasmic side of vesicle fuses with cytoplasmic side of plasma membrane. Thus, the contents of vesicles are externalized. This process is called exocytosis. Release of trypsinogen by pancreatic acinar cells and release of insulin by beta cells of Langerhans are examples of exocytosis. Often, hormones or changes in calcium ion concentration are the signal for exocytosis.

Endocytosis

Endocytosis is the mechanism by which cells internalize extracellular macromolecules, to form an endocytic vesicle. This requires energy in the form of ATP as well as calcium ions in the extracellular fluid. Cytoplasmic contractile elements take part in this movement. In general, plasma membrane is invaginated, enclosing the matter. This forms the endocytic

vesicle. The endocytosis may be phagocytosis or receptor

mediated endocytosis.

Receptor Mediated Endocytosis

Low density lipoprotein (LDL) binds to the LDL receptor and the complex is later internalized. The cytoplasmic side of these vesicles are coated with filaments; mainly composed of Clathrin. These are called Clathrin coated pits. Several hormones are also taken up by the cells by receptor-mediated mechanism. Many viruses get attached to their specific receptors on the cell membranes. Examples are influenza virus, hepatitis B virus, poliovirus and HIV.

Phagocytosis

The term is derived from the Greek word "phagein" which means to eat. It is the engulfment of large particles such as bacteria by

macrophages and granulocytes. They extend pseudopodia

and surround the particles to form phagosomes. Phagosomes later fuse with lysosomes to form phagolysosomes, inside which the particles are digested. An active macrophage can ingest 25% of their volume per hour.

A QUICK LOOK

• Cell is the basic unit of all living organisms. • In a cell, biomolecules are maintained in a state

of ‘dynamic’ or ‘steady state’ equilibrium.

• All cells in the body contain nucleus except mature erythrocytes.

• Endoplasmic reticulum is involved in protein synthesis and also detoxification of various drugs. • Golgi apparatus is primarily involved in glycosy-lation, protein sorting, packaging and secretion. • Lysosom es are packets containing hydrolytic

enzymes.

• Mitochondria are the ‘power house’ of the cell. They have their own DNA. They can synthesize their own proteins.

• Antibiotics inhibiting bacterial protein biosynthesis can inhibit mitochondrial protein biosynthesis also. • Mem branes are m ainly com posed of l ipids (phospholipids), proteins and a small percentage of carbohydrates.

• Phospholipids, which are amphipathic in nature, are arranged as bilayers.

• Cholesterol content and nature of the fatty acid of the membrane, influence the fluidity.

• Membrane proteins can be integral, peripheral or transmembrane.

• Transmembrane proteins serve as receptors,

tissue specific antigens, ion-channels, etc. • Transport of m ol ecules across the pl asm a

membrane could be energy dependent (Active) or energy independent (Passive).

• Ion-channels function for the transport of the ions such as Ca2+, K+, Cl, Na+, etc.

• Na+-K+ ATPase (Sodium Pump) is an example of

Active transport. Cardiotonic drugs like Digoxin and Ouabain competitively inhibit K+ ion binding.

The property is used to enhance contractility of the cardiac muscle.

• Transport systems may be Uniport, Antiport or Symport.

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CHAPTER AT A GLANCE

The reader will be able to answer questions on the following topics:

1. Classification of amino acids based on structure 2. Based on side chain character

3. Based on metabolic fate

4. Based on nutritional requirements 5. Isoelectric point

6. Reactions due to carboxyl and amino groups 7. Peptide bond formation

8. Primary, secondary, tertiary and quaternary structure of proteins

9. Precipitation reactions of proteins 10. Classification of proteins

The word protein is derived from Greek word, “proteios” which means primary. As the name shows, the proteins are of paramount importance for biological systems. Out of the total dry body weight, 3/4ths are made up of proteins. Proteins are used for body building ; all the major structural and functional aspects of the body are carried out by protein molecules. Abnormality in protein structure will lead to molecular diseases with profound alterations in metabolic functions.

Proteins contain Carbon, Hydrogen, Oxygen and Nitrogen as the major components while Sulphur and Phosphorus are minor constituents. Nitrogen is characteristic of proteins. On an average, the

nitrogen content of ordinary proteins is 16% by weight.

All proteins are polymers of amino acids. Commonly occurring amino acids are 20 in number. Most of the amino acids (except proline) are alpha amino acids, which means that the amino group is attached to the same carbon atom to which the carboxyl group is attached (Fig. 2.1).

CLASSIFICATION OF AMINO ACIDS

1. Based on Structure A. Aliphatic Amino Acids

a. Mono amino mono carboxylic acids: • Simple amino acids:

1. Glycine, 2. Alanine (Fig. 2.2)

Amino Acids and Proteins

Fig. 2.2: Simple amino acids • Branched chain amino acids:

3. Valine, 4. Leucine, 5. Isoleucine (Fig. 2.3)

• Hydroxy amino acids:

6. Serine, 7. Threonine (Fig. 2.4) • Sulfur containing:

8. Cysteine, 9. Methionine (Fig. 2.5)

Fig. 2.1: General

structure

Fig. 2.3: Branched chain amino acids

Fig. 2.4: Hydroxy amino acids

Fig. 2.5: Sulfur

containing amino acids

2

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Fig. 2.12: Some derived amino acids

proline (Fig. 2.12) and hydroxy lysine are important components of collagen. Gamma carboxylation of glutamic acid residues of proteins is important for clotting process (Fig. 2.12).

b. Derived amino acids not seen in proteins: (Non-protein amino acids): Some derived amino acids are seen free in cells, e.g. Ornithine (see Chapter 12), Citrulline, Homocysteine. These

Fig. 2.9: Aromatic amino acids

Fig. 2.8: Dibasic amino acids

• Having amide group:

10. Asparagine, 11. Glutamine (Fig. 2.6) b. Mono amino dicarboxylic acids (Fig. 2.7):

12. Aspartic acid, 13. Glutamic acid c. Dibasic mono carboxylic acids:

14. Lysine, 15. Arginine (Fig. 2.8)

B. Aromatic Amino Acids

16. Phenylalanine, 17. Tyrosine (Fig. 2.9)

C. Heterocyclic Amino Acids

18. Tryptophan (Fig. 2.10), 19. Histidine (Fig. 2.11)

Fig. 2.6: Amino acids

with amide groups

Fig. 2.7: Dicarboxylic

amino acids

D. Imino Acid

20. Proline (Fig. 2.11)

E. Derived Amino Acids

a. Derived amino acids found in proteins: After the synthesis of proteins, some of the amino acids are modified, e.g. hydroxy

Fig. 2.10: Tryptophan (Trp) (W) with indole group

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are produced during the metabolism of amino acids.

Each amino acid will have three-letter and one letter abbreviations which are shown in Figures 2.2 to 2.11, as well as in Table 2.1.

Special Groups in Amino Acids

Arginine contains guanidinium group (-NH-CNH— NH2); Phenylalanine contains benzene group; Tyrosine (phenol); Tryptophan (Indole group); Histidine (imidazole); and Proline (pyrrolidine) (Table 2.1). Proline has a secondary amino group, and hence it is an imino acid.

2. Classification Based on Side Chain

A. Amino Acids having Nonpolar Side Chains

These include Alanine, Valine, Leucine, Isoleucine, Methionine, Proline, Phenylalanine and Tryptophan. These groups are hydrophobic (water repellant) and lipophilic.

B. Amino Acids having Uncharged or Nonionic Polar Side Chains

Glycine, Serine, Threonine, Cysteine, Tyrosine, Glutamine and Asparagine belong to this group. These amino acids are hydrophilic in nature.

C. Amino Acids having Charged or Ionic Polar Side Chains

They are hydrophilic in nature.

a. Acidic amino acids: They have a negative charge on the R group: Aspartic acid and Glutamic acid. (Tyrosine is mildly acidic). b. Basic amino acids: They have a positive

charge on the R group: Lysine, Arginine and Histidine.

3. Classification Based on Metabolic Fate A. Purely Ketogenic

Leucine is purely ketogenic because it will enter into the metabolic pathway of ketogenesis.

B. Ketogenic and Glucogenic

Lysine, Isoleucine, Phenylalanine, Tyrosine and Tryptophan are partially ketogenic and partially glucogenic. During metabolism, part of the carbon skeleton of these amino acids will enter the fatty acid metabolic pathway and the other part into glucose pathway.

C. Purely Glucogenic

All the remaining 14 amino acids are purely glucogenic as they enter only into the glucogenic pathway.

4. Classification Based on Nutritional Requirement

A. Essential or Indispensable

The amino acids may further be classified according to their essentiality for growth. Thus, Isoleucine, Leucine, Threonine, Lysine, Methionine, Phenyl-alanine, Tryptophan, and Valine are essential amino acids. Their carbon skeleton cannot be synthesized by human beings and so preformed amino acids are to be taken in food for normal growth. One tip to remember these names is given in Box 2.1.

B. Partially Essential or Semi-essential

Histidine and Arginine are semi-indispensable amino acids. Growing children require them in food. But they are not essential for the adult individual.

C. Nonessential or Dispensable

The remaining 10 amino acids are nonessential. However, they are also required for the normal Table 2.1: Common amino acids

Name of Special 3-

1-amino acid group letter letter present abbre. abbre .

Glycine Gly G

Alanine Ala A

Valine Val V

Leucine Leu L

Isoleucine Ile I

Serine Hydroxyl Ser S

Threonine Hydroxyl Thr T

Cysteine Sulfhydryl Cys C

Methionine Thioether Met M

Asparagine Amide Asn N

Glutamine Amide Gln Q

Aspartic acid Beta-carboxyl Asp D

Glutamic acid Gamma-carboxyl Glu E

Lysine Epsilon-amino Lys K

Arginine Guanidinium Arg R

Phenylalanine Benzene Phe F

Tyrosine Phenol Tyr Y

Tryptophan Indole Trp W

Histidine Imidazole His H

References

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