Abstract
Current limitations in de novo protein structure prediction and design suggest an incomplete understanding of the interactions that govern protein folding. Here we demonstrate that previously unappreciated hydrogen bonds occur within proteins between the amide proton and carbonyl oxygen of the same residue. Quantum calculations, infrared spectroscopy, and nuclear magnetic resonance spectroscopy show that these interactions share hallmark features of canonical hydrogen bonds. Biophysical analyses demonstrate that selective attenuation or enhancement of these C5 hydrogen bonds affects the stability of synthetic β-sheets. These interactions are common, affecting approximately 5% of all residues and 94% of proteins, and their cumulative impact provides several kilocalories per mole of conformational stability to a typical protein. C5 hydrogen bonds especially stabilize the flat β-sheets of the amyloid state, which is linked with Alzheimer's disease and other neurodegenerative disorders. Inclusion of these interactions in computational force fields would improve models of protein folding, function, and dysfunction.
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References
Dill, K.A. & MacCallum, J.L. The protein-folding problem, 50 years on. Science 338, 1042–1046 (2012).
Dill, K.A. Dominant forces in protein folding. Biochemistry 29, 7133–7155 (1990).
Pauling, L., Corey, R.B. & Branson, H.R. The structure of proteins; two hydrogen-bonded helical configurations of the polypeptide chain. Proc. Natl. Acad. Sci. USA 37, 205–211 (1951).
Pauling, L. & Corey, R.B. The pleated sheet, a new layer configuration of polypeptide chains. Proc. Natl. Acad. Sci. USA 37, 251–256 (1951).
Kabsch, W. & Sander, C. Dictionary of protein secondary structure: pattern recognition of hydrogen-bonded and geometrical features. Biopolymers 22, 2577–2637 (1983).
Clauss, A.D. et al. Rabbit-ears hybrids, VSEPR sterics, and other orbital anachronisms. Chem. Educ. Res. Pract. 15, 417–434 (2014).10.1039/C4RP00057A
Bartlett, G.J., Choudhary, A., Raines, R.T. & Woolfson, D.N. n→π* interactions in proteins. Nat. Chem. Biol. 6, 615–620 (2010).
Bartlett, G.J. & Woolfson, D.N. On the satisfaction of backbone-carbonyl lone pairs of electrons in protein structures. Protein Sci. 25, 887–897 (2016).10.1002/pro.2896
Avignon, M., Huong, P.V., Lascombe, J., Marraud, M. & Neel, J. Etude, par spectroscopie infra-rouge, de la conformation de quelques composés peptidiques modèles. Biopolymers 8, 69–89 (1969).
Burgess, A.W. & Scheraga, H.A. Communications to the editor: Stable conformations of dipeptides. Biopolymers 12, 2177–2183 (1973).
Toniolo, C. Intramolecularly hydrogen-bonded peptide conformations. CRC Crit. Rev. Biochem. 9, 1–44 (1980).
Dian, B.C. et al. The infrared and ultraviolet spectra of single conformations of methyl-capped dipeptides: N-acetyl tryptophan amide and N-acetyl tryptophan methyl amide. J. Chem. Phys. 117, 10688–10702 (2002).
Blanco, S., Lesarri, A., López, J.C. & Alonso, J.L. The gas-phase structure of alanine. J. Am. Chem. Soc. 126, 11675–11683 (2004).10.1021/ja048317c
González Flórez, A.I. et al. Charge-induced unzipping of isolated proteins to a defined secondary structure. Angew. Chem. Int. Ed. Engl. 55, 3295–3299 (2016).10.1002/anie.201510983
Zimmerman, S.S., Pottle, M.S., Némethy, G. & Scheraga, H.A. Conformational analysis of the 20 naturally occurring amino acid residues using ECEPP. Macromolecules 10, 1–9 (1977).
Scheiner, S. Relative strengths of NH.O and CH.O hydrogen bonds between polypeptide chain segments. J. Phys. Chem. B 109, 16132–16141 (2005).10.1021/jp053416d
Steiner, T. The hydrogen bond in the solid state. Angew. Chem. Int. Ed. Engl. 41, 48–76 (2002).
McDonald, I.K. & Thornton, J.M. Satisfying hydrogen bonding potential in proteins. J. Mol. Biol. 238, 777–793 (1994).
Rowland, R.S. & Taylor, R. Intermolecular nonbonded contact distances in organic crystal structures: comparison with distances expected from van der Waals radii. J. Phys. Chem. 100, 7384–7391 (1996).10.1021/jp953141
Reed, A.E., Curtiss, L.A. & Weinhold, F. Intermolecular interactions from a natural bond orbital, donor–acceptor viewpoint. Chem. Rev. 88, 899–926 (1988).
Hinderaker, M.P. & Raines, R.T. An electronic effect on protein structure. Protein Sci. 12, 1188–1194 (2003).
Choudhary, A., Gandla, D., Krow, G.R. & Raines, R.T. Nature of amide carbonyl–carbonyl interactions in proteins. J. Am. Chem. Soc. 131, 7244–7246 (2009).
Newberry, R.W., VanVeller, B., Guzei, I.A. & Raines, R.T. n→π* interactions of amides and thioamides: implications for protein stability. J. Am. Chem. Soc. 135, 7843–7846 (2013).10.1021/ja4033583
Benedetti, E. et al. Structural versatility of peptides from Cα,α-dialkylated glycines. I. A conformational energy computation and X-ray diffraction study of homo-peptides from Cα,α-diethylglycine. Biopolymers 27, 357–371 (1988).10.1002/bip.360270302
Toniolo, C. et al. Structural versatility of peptides from Cα,α-dialkylated glycines. II. An IR absorption and 1H-NMR study of homo-oligopeptides from Cα,α-diethylglycine. Biopolymers 27, 373–379 (1988).10.1002/bip.360270303
Deechongkit, S. et al. Context-dependent contributions of backbone hydrogen bonding to β-sheet folding energetics. Nature 430, 101–105 (2004).
Dado, G.P. & Gellman, S.H. Structural and thermodynamic characterization of temperature-dependent changes in the folding pattern of a synthetic triamide. J. Am. Chem. Soc. 115, 4228–4245 (1993).10.1021/ja00063a046
Skinner, J.J., Lim, W.K., Bédard, S., Black, B.E. & Englander, S.W. Protein dynamics viewed by hydrogen exchange. Protein Sci. 21, 996–1005 (2012).10.1002/pro.2081
Cochran, A.G., Skelton, N.J. & Starovasnik, M.A. Tryptophan zippers: stable, monomeric β-hairpins. Proc. Natl. Acad. Sci. USA 98, 5578–5583 (2001).
Fu, Y., Gao, J., Bieschke, J., Dendle, M.A. & Kelly, J.W. Amide-to-E-olefin versus amide-to-ester backbone H-bond perturbations: evaluating the O-O repulsion for extracting H-bond energies. J. Am. Chem. Soc. 128, 15948–15949 (2006).
Derewenda, Z.S., Lee, L. & Derewenda, U. The occurrence of C-H...O hydrogen bonds in proteins. J. Mol. Biol. 252, 248–262 (1995).
Culik, R.M., Jo, H., DeGrado, W.F. & Gai, F. Using thioamides to site-specifically interrogate the dynamics of hydrogen bond formation in β-sheet folding. J. Am. Chem. Soc. 134, 8026–8029 (2012).10.1021/ja301681v
Gallivan, J.P. & Dougherty, D.A. Cation-π interactions in structural biology. Proc. Natl. Acad. Sci. USA 96, 9459–9464 (1999).
Shi, Z., Chen, K., Liu, Z. & Kallenbach, N.R. Conformation of the backbone in unfolded proteins. Chem. Rev. 106, 1877–1897 (2006).
Richardson, J.S. & Richardson, D.C. Natural β-sheet proteins use negative design to avoid edge-to-edge aggregation. Proc. Natl. Acad. Sci. USA 99, 2754–2759 (2002).
Cheng, P.-N., Pham, J.D. & Nowick, J.S. The supramolecular chemistry of β-sheets. J. Am. Chem. Soc. 135, 5477–5492 (2013).
Sawaya, M.R. et al. Atomic structures of amyloid cross-β spines reveal varied steric zippers. Nature 447, 453–457 (2007).
Dobson, C.M. Protein misfolding, evolution and disease. Trends Biochem. Sci. 24, 329–332 (1999).
Ohnishi, S., Kamikubo, H., Onitsuka, M., Kataoka, M. & Shortle, D. Conformational preference of polyglycine in solution to elongated structure. J. Am. Chem. Soc. 128, 16338–16344 (2006).10.1021/ja066008b
Minor, D.L. Jr. & Kim, P.S. Measurement of the β-sheet-forming propensities of amino acids. Nature 367, 660–663 (1994).
Kortemme, T., Morozov, A.V. & Baker, D. An orientation-dependent hydrogen bonding potential improves prediction of specificity and structure for proteins and protein-protein complexes. J. Mol. Biol. 326, 1239–1259 (2003).
Reed, A.E., Weinstock, R.B. & Weinhold, F. Natural population analysis. J. Chem. Phys. 83, 735–746 (1985).
Huang, R. et al. Cross-strand coupling and site-specific unfolding thermodynamics of a trpzip β-hairpin peptide using 13C isotopic labeling and IR spectroscopy. J. Phys. Chem. B 113, 5661–5674 (2009).10.1021/jp9014299
Wang, G. & Dunbrack, R.L. Jr. PISCES: a protein sequence culling server. Bioinformatics 19, 1589–1591 (2003).
Hutchinson, E.G. & Thornton, J.M. PROMOTIF--a program to identify and analyze structural motifs in proteins. Protein Sci. 5, 212–220 (1996).
Acknowledgements
We thank G.J. Bartlett, I.C. Tanrikulu, and L.L. Kiessling for discussions, and W.M. Westler, T. Zhang, and M.T. Zanni for assistance with spectroscopy. This work was supported by grants R01 AR044276 (NIH), R01 GM044783 (NIH), and CHE-1124944 (NSF). R.W.N. was supported by Biotechnology Training Grant T32 GM008349 (NIH) and by an ACS Division of Organic Chemistry Graduate Fellowship. The National Magnetic Resonance Facility at Madison is supported by grant P41 GM103399 (NIH). High-performance computing is supported by grant CHE-0840494 (NSF). The Biophysics Instrumentation Facility at the University of Wisconsin–Madison was established with grants BIR-9512577 (NSF) and S10 RR013790 (NIH).
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R.W.N. conceived of the project. R.W.N. and R.T.R. planned the experiments. R.W.N. carried out the experiments. R.W.N. and R.T.R. analyzed the data and wrote the manuscript.
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Supplementary Results, Supplementary Tables 1–6, Supplementary Figures 1–13 and Supplementary Note 1. (PDF 2464 kb)
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Synthetic Procedures. (PDF 3388 kb)
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Newberry, R., Raines, R. A prevalent intraresidue hydrogen bond stabilizes proteins. Nat Chem Biol 12, 1084–1088 (2016). https://doi.org/10.1038/nchembio.2206
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DOI: https://doi.org/10.1038/nchembio.2206
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