determination of protein stoichiometry
5. Conclusions and Future Prospects
Φ6 PC assembly is precise and efficient. Electrostatic interactions are the main force driving Φ6 PC formation (III). P4 hexamers induce the nucleation of PC assembly (Poranen et al., 2001), and initially associate to each of the PC’s five-fold vertices (I). Proteins P7 and P2 share similar binding positions within the PC shell (Katz et al., 2012; Nemecek et al., 2012; Sen et al., 2008) and may interact during the PC assembly process (I). Although P4-deficient PCs may undergo premature expansion, these particles can efficiently sequester free P4 hexamer from the environment to recover their normal compact conformation and associated capacity for RNA encapsidation and transcription (II).
Many issues remain to be studied. For instance, in vitro systems of Φ6 PC self- assembly and encapsidation can be used to detect the proposed assembly and packaging intermediates. Highly uniform, self-assembled particles with full P2 or P7 occupancy provide opportunities to define the precise interactions of these proteins with the PC shell in high-resolution structural studies. Furthermore, the position of the P2 polymerase within the PC during particle maturation is an open question: does it stay in the three-fold symmetry position or move toward the five-fold vertex? In addition, how P2 activity is associated with PC conformational switches remains to be investigated. Comparison of wt PCs and P4-deficient particles may provide clues to the location of P2 in expanded particles.
6. Acknowledgements
This work was carried out at the Academy of Finland Centre of Excellence in Virus Research (2006-2011) and the Programme on Molecular Virology, in the Department of Biosciences and Institute of Biotechnology, at the University of Helsinki, under the supervision of Professor Dennis Bamford and Docent Minna Poranen.
I would like to thank my dear supervisors Minna and Dennis. Thanks to Dennis for providing me the opportunity to work in the DB-lab. I appreciate your wisdom, enthusiasm and support to my thesis work. I wish to express my deepest gratitude to Minna, for the constant encourage, trust and patience. Thank you Minna for guiding me to the fantastic world of viral self-assembly, for every discussion we had and for always giving useful suggestions to solve any problem. I would like to thank Minna also for leaving a table for me next to her and answering my numerous questions about science and life.
I would like to thank Professor Albie van Dijk and Dr. Denis Kainov for carefully reviewing my dissertation and giving excellent comments within such a short period. I thank Docent Kristiina Mäkinen and Dr. Denis Kainov for being the members of my thesis committee. I appreciate all the valuable advices and critical comments from you on my thesis work. I am grateful to Docent Sarah Butcher for giving advices on my study and work.
The Viikki Doctoral Programme in Molecular Biosciences (VGSB) is acknowledged for the financial support to my study and conference trips, as well as providing interesting courses.
I am grateful to my co-author, Markus Pirttimaa, for his brilliant ideas and valuable comments on the manuscript. I would like to thank all the former and current members in the DB-lab for creating the lovely atmosphere. I am grateful to Antti, Miao, Linda, Gabija, Virginija, Heli, Janne, Nina, Ana, Alesia, Maija, Peter and Julija for the friendship and support. Thanks to Antti for teaching me all the basic techniques of gene cloning and protein purifications, as well as guiding me to join the DB-lab lunch break. Thanks to Miao for the endless discussion about work, life and future, as well as nice food offered by you and your wife Lijing. Thanks to Heli for the company in the conferences and taking care of me in Switzerland. Thanks to Janne for helping me to solve many computer problems and introducing the dentist to me.
I would like to thank our highly skilled technicians, especially for preparing hundreds of SDS-polyacrylamide gels used in my thesis work. I deeply thank Riitta for all the help and friendship. I appreciate your excellent technical assistance and optimism. It is always enjoyable to work with you.
I would like to thank my dear friends, Yurui Tang, Fang Wang, Ping Jiang, Li Ma and Lin Ning, for all the moments we spent together throughout the years. Those memories of happiness and tears are my treasure. Thanks to Yurui for being my ‘classmate’ during my postgraduate studies, for going to library together and sharing dinner when working late. Thanks to Xing, Shentong, Chaoyi, Xin, Ying, Ruiqing, Zhao, Lin and all others for the friendship.
I would like to thank my best friend Zhongwen for his support and care, and all the gift packages sent from China every several months. I am deeply grateful to my aunt family, Sanwei, Wangning, Yingzi and Bin, for all the help since I came to Finland and for the great time we shared in your sweet home. Lastly, I would like to thank my mother Yuan for her constant love, support, care and encouragement. Thank you mom for being a model for me of a nice person and a good scientist.
Helsinki 2014-04
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