Design method for targeting PCSK9 - Supporting Information

2.5 Supporting Information

3.5.1 Design method for targeting PCSK9

The following is the RosettaScripts method used to design scaffold proteins to bind to PCSK9 rosetta3/rosetta_source/bin/rosetta_scripts.<EXE> -database <ROSETTA_DATABASE> -s input_struct.pdb -ignore_unrecognized_res -parser:protocol script.xml

-nstruct 9000 -jd2:ntrials 5 -jd2:delete_old_poses -out:pdb_gz true

-score12prime true -no_his_his_pairE -no_optH false -docking:dock_pert 3 5

-ex1 -ex2 -use_input_sc -extrachi_cutoff 1 -linmem_ig 10

-run::min_type dfpmin_armijo_nonmonotone

-atomic_burial_cutoff 0.01 -sasa_calculator_probe_radius 1.2 -overwrite

With the following as the input toscript.xml.

< R O S E T T A S C R I P T S > < S C O R E F X N S > < s 1 2 _ p r i m e w e i g h t s = " s c o r e 1 2 p r i m e " / > < / S C O R E F X N S > < T A S K O P E R A T I O N S > < R e s t r i c t T o I n t e r f a c e V e c t o r n a m e = v e c t o r T a s k c h a i n 1 _ n u m =1 c h a i n 2 _ n u m =2 C B _ d i s t _ c u t o f f = 1 0 . 0 n e a r b y _ a t o m _ c u t o f f = 5 . 5 v e c t o r _ a n g l e _ c u t o f f = 6 5 . 0 v e c t o r _ d i s t _ c u t o f f = 8 . 0 / > < R e s t r i c t C h a i n T o R e p a c k i n g n a m e = r e p a c k 1 c h a i n =1/ > < R e s t r i c t C h a i n T o R e p a c k i n g n a m e = r e p a c k 2 c h a i n =2/ > < R e s t r i c t T o R e p a c k i n g n a m e = r e p a c k o n l y / > < / T A S K O P E R A T I O N S > < F I L T E R S > < S c o r e T y p e n a m e = s c o r e _ f i l t e r s c o r e f x n = s c o r e 1 2 s c o r e _ t y p e = t o t a l _ s c o r e t h r e s h o l d = -1/ > < Ddg n a m e = d d g _ f i l t e r s c o r e f x n = s c o r e 1 2 t h r e s h o l d = -10 j u m p =1 r e p e a t s =3 r e p a c k =1/ > < S a s a n a m e = s a s a _ f i l t e r t h r e s h o l d = 9 0 0 j u m p =1/ > < R m s d n a m e = rms t h r e s h o l d = 5 . 0 c o n f i d e n c e =1/ > < / F I L T E R S > < M O V E R S > < M i n P a c k M o v e r n a m e = d e s i g n _ m i n _ p a c k s c o r e f x n = s c o r e 1 2 t a s k _ o p e r a t i o n s = v e c t o r T a s k , r e p a c k 1 / > 70

< P r e p a c k n a m e = p r e p a c k s c o r e f x n = s 1 2 _ p r i m e j u m p _ n u m b e r =1 m i n _ b b =0 t a s k _ o p e r a t i o n s = v e c t o r T a s k , r e p a c k o n l y , cmdTask , c u r r e n t T a s k / > < D o c k i n g P r o t o c o l n a m e = d o c k p r o t o c o l d o c k i n g _ l o c a l _ r e f i n e =0

d o c k i n g _ s c o r e _ h i g h = s c o r e 1 2 i g n o r e _ d e f a u l t _ d o c k i n g _ t a s k =1

t a s k _ o p e r a t i o n s = v e c t o r T a s k , repack1 , repack2 , cmdTask , c u r r e n t T a s k / > < T a s k A w a r e M i n M o v e r n a m e = m i n m o v e r s c o r e f x n = s 1 2 _ p r i m e chi =1 bb =1 j u m p =1 t a s k _ o p e r a t i o n s = cmdTask , c u r r e n t T a s k / > Add m o v e r _ n a m e = d e s i g n _ m i n _ p a c k < Add m o v e r _ n a m e = d e s i g n _ p a c k / > < Add m o v e r _ n a m e = m i n m o v e r / > < S a v e A n d R e t r i e v e S i d e c h a i n s n a m e = r e t r i e v e _ s i d e c h a i n s a l l s c =0/ > < I n t e r f a c e A n a l y z e r M o v e r n a m e = f u l l a n a l y z e s c o r e f x n = s c o r e 1 2 p a c k s t a t =1 p a c k _ i n p u t =0 j u m p =1 t r a c e r =0 u s e _ j o b n a m e =1 r e s f i l e =0 / > < F a v o r N a t i v e R e s i d u e n a m e = f a v o r n a t i v e b o n u s = 0 . 5 / > < / M O V E R S > < A P P L Y _ T O _ P O S E > < / A P P L Y _ T O _ P O S E > < Add m o v e r _ n a m e = p r e p a c k / > Add m o v e r _ n a m e = a l a _ p o s e Add m o v e r _ n a m e = f a v o r n a t i v e < Add m o v e r _ n a m e = d o c k p r o t o c o l f i l t e r _ n a m e = rms / > Add m o v e r _ n a m e = r e t r i e v e _ s i d e c h a i n s < Add m o v e r _ n a m e = d e s i g n i t e r a t e f i l t e r _ n a m e = rms / > < Add f i l t e r _ n a m e = s c o r e _ f i l t e r / > < Add f i l t e r _ n a m e = d d g _ f i l t e r / > < Add f i l t e r _ n a m e = s a s a _ f i l t e r / > < Add f i l t e r _ n a m e = u n s a t s _ f i l t e r / > < Add m o v e r _ n a m e = f u l l a n a l y z e / > < / R O S E T T A S C R I P T S >

References

1. Stranges, P. B., Machius, M., Miley, M. J., Tripathy, A., and Kuhlman, B. (2011) Com- putational design of a symmetric homodimer using β-strand assembly. Proceedings of the National Academy of Sciences of the United States of America108, 20562–7

2. Der, B. S., Machius, M., Miley, M. J., Mills, J. L., Szyperski, T., and Kuhlman, B. (2012) Metal-Mediated Affinity and Orientation Specificity in a Computationally Designed Protein Homodimer. Journal of the American Chemical Society 134, 375–385

3. Fleishman, S. J., Whitehead, T. A., Ekiert, D. C., Dreyfus, C., Corn, J. E., Strauch, E.- M. M., Wilson, I. A., and Baker, D. (2011) Computational design of proteins targeting the conserved stem region of influenza hemagglutinin. Science 332, 816–821

4. Der, B. S. and Kuhlman, B. (2011) Biochemistry. From computational design to a protein that binds. Science332, 801–2

5. Bodemann, B. O. and White, M. A. (2008) Ral GTPases and cancer: linchpin support of the tumorigenic platform. Nature Reviews Cancer8, 133–140

6. Aguirre-Ghiso, J. A., Frankel, P., Farias, E. F., Lu, Z. M., Jiang, H., Olsen, A., Feig, L. A., Joffe, E. B., and Foster, D. A. (1999) RalA requirement for v-Src- and v-Ras- induced tumorigenicity and overproduction of urokinase-type plasminogen activator: involvement of metalloproteases. Oncogene18, 4718–4725

7. Urano, T., Emkey, R., and Feig, L. A. (1996) Ral-GTPases mediate a distinct down- stream signaling pathway from Ras that facilitates cellular transformation. Embo Journal15, 810–816

8. Fukai, S., Matern, H. T., Jagath, J. R., Scheller, R. H., and Brunger, A. T. (2003) Structural basis of the interaction between RalA and Sec5, a subunit of the sec6/8 complex. The EMBO journal 22, 3267–78

9. Jin, R., Junutula, J., Matern, H., Ervin, K., Scheller, R., and Brunger, A. (2005) Exo84 and Sec5 are competitive regulatory Sec6/8 effectors to the RalA GTPase. EMBO J

24, 2064–2074

10. Sheinerman, F. B. and Honig, B. (2002) On the role of electrostatic interactions in the design of protein-protein interfaces. Journal Of Molecular Biology318, 161–177

11. Kiel, C., Serrano, L., and Herrmann, C. (2004) A detailed thermodynamic analysis of Ras/effector complex interfaces. Journal Of Molecular Biology 340, 1039–1058

12. Nassar, N., Horn, G., Herrmann, C., Block, C., Janknecht, R., and Wittinghofer, A. (1996) Ras/Rap effector specificity determined by charge reversal. Nature structural biology 3, 723–9

13. Kiel, C., Selzer, T., Shaul, Y., Schreiber, G., and Herrmann, C. (2004) Electrostatically optimized Ras-binding Ral guanine dissociation stimulator mutants increase the rate of association by stabilizing the encounter complex. Proceedings of the National Academy of Sciences of the United States of America101, 9223–8

14. Filchtinski, D., Sharabi, O., R¨uppel, A., Vetter, I. R., Herrmann, C., and Shifman, J. M. (2010) What makes Ras an efficient molecular switch: a computational, bio- physical, and structural study of Ras-GDP interactions with mutants of Raf. Journal of molecular biology399, 422–35

15. Horton, J. D., Cohen, J. C., and Hobbs, H. H. (2007) Molecular biology of PCSK9: its role in LDL metabolism. Trends in biochemical sciences32, 71–7

16. Lagace, T. A., Curtis, D. E., Garuti, R., McNutt, M. C., Park, S. W., Prather, H. B., Anderson, N. N., Ho, Y. K., Hammer, R. E., and Horton, J. D. (2006) Secreted PCSK9 decreases the number of LDL receptors in hepatocytes and in livers of parabiotic mice. The Journal of clinical investigation116, 2995–3005

17. Kwon, H. J., Lagace, T. A., McNutt, M. C., Horton, J. D., and Deisenhofer, J. (2008) Molecular basis for LDL receptor recognition by PCSK9. Proceedings of the National Academy of Sciences of the United States of America105, 1820–5

18. Cohen, J. C., Boerwinkle, E., Mosley, T. H., and Hobbs, H. H. (2006) Sequence variations in PCSK9, low LDL, and protection against coronary heart disease. The New England journal of medicine 354, 1264–72

19. Abifadel, M., Varret, M., Rabès, J.-P., Allard, D., Ouguerram, K., Devillers, M., Cru- aud, C., Benjannet, S., Wickham, L., Erlich, D., Derré, A., Villéger, L., Farnier, M., Beucler, I., Bruckert, E., Chambaz, J., Chanu, B., Lecerf, J.-M., Luc, G., Moulin, P., Weissenbach, J., Prat, A., Krempf, M., Junien, C., Seidah, N. G., and Boileau, C. (2003) Mutations in PCSK9 cause autosomal dominant hypercholesterolemia. Nature

genetics 34, 154–6

20. Ni, Y. G., Di Marco, S., Condra, J. H., Peterson, L. B., Wang, W., Wang, F., Pandit, S., Hammond, H. A., Rosa, R., Cummings, R. T., Wood, D. D., Liu, X., Bottomley, M. J., Shen, X., Cubbon, R. M., Wang, S.-p., Johns, D. G., Volpari, C., Hamuro, L., Chin, J., Huang, L., Zhao, J. Z., Vitelli, S., Haytko, P., Wisniewski, D., Mitnaul, L. J., Sparrow, C. P., Hubbard, B., Carf´ı, A., and Sitlani, A. (2011) A PCSK9-binding antibody that structurally mimics the EGF(A) domain of LDL-receptor reduces LDL cholesterol in vivo. Journal of lipid research52, 78–86

21. Orcutt, K. D., Wittrup, K. D., Kontermann, R., and D¨ubel, S. (2010) Antibody Engineering. Springer Berlin Heidelberg, Berlin, Heidelberg. ISBN 978- 3-642-01143-6. doi:10.1007/978-3-642-01144-3

22. Gai, S. A. and Wittrup, K. D. (2007) Yeast surface display for protein engineering and characterization. Current opinion in structural biology17, 467–73

23. Chen, I., Dorr, B. M., and Liu, D. R. (2011) A general strategy for the evolution of bond-forming enzymes using yeast display. Proceedings of the National Academy of Sciences of the United States of America108, 11399–404

24. Chao, G., Lau, W. L., Hackel, B. J., Sazinsky, S. L., Lippow, S. M., and Wittrup, K. D. (2006) Isolating and engineering human antibodies using yeast surface display. Nature

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