Abstract
A simple method for measuring amide hydrogen exchange rates is presented, which is based on the selective inversion of water magnetization with the use of radiation damping. Simulations show that accurate exchange rates can be measured despite the complications of radiation damping and cross relaxation to the exchange process between amide and water protons. This method cannot eliminate the contributions of the exchange-relayed NOE and direct NOE to the measured exchange rates, but minimize the direct NOE contribution. In addition, the amides with a significant amount of such indirect contributions are possible to be identified from the shape of the exchange peak intensity profiles or/and from the apparent relaxation rates of amide protons which are extracted from fitting the intensity profiles to an equation established here for our experiment. The method was tested on ubiquitin and also applied to an acyl carrier protein. The amide exchange rates for the acyl carrier protein at two pHs indicate that the entire protein is highly dynamic on the second timescale. Low protection factors for the residues in the regular secondary structural elements also suggest the presence of invisible unfolded species. The highly dynamic nature of the acyl carrier protein may be crucial for its interactions with its substrate and enzymes.
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References
Andrec M, Hill RB, Prestegard JH (1995) Amide exchange rates in Escherichia coli acyl carrier protein: Correlation with protein structure and dynamics. Protein Sci 4:983–993
Bai YW, Milne JS, Mayne L, Englander SW (1993) Primary structure effects on peptide group hydrogen exchange. Proteins Struct Funct Genet 17:75–86
Bockmann A, Penin F, Guittet E (1996) Rapid estimation of relative amide proton exchange rates of 15 N-labelled proteins by a straightforward water selective NOESY-HSQC experiment. FEBS Lett 383:191–195
Chan DI, Vogel HJ (2010) Current understanding of fatty acid biosynthesis and the acyl carrier protein. Biochem J 430:1–19
Chen J-H, Cutting B, Bodenhausen G (2000) Measurement of radiation damping rate constants in nuclear magnetic resonance by inversion recovery and automated compensation of selective pulses. J Chem Phys 112:6511–6514
Chevelkov V, Xue Y, Rao DK, Forman-Kay JD, Skrynnikov NR (2010) 15 N H/D-SOLEXSY experiment for accurate measurement of amide solvent exchange rates: application to denatured drkN SH3. J Biomol NMR 46:227–244
Christoffersen M, Bolvig S, Tüchsen E (1996) Salt effects on the amide hydrogen exchange of bovine pancreatic trypsin inhibitor. Biochemistry 35:2309–2315
Connelly GP, Bai Y, Jeng MF, Englander SW (1993) Isotope effects in peptide group hydrogen exchange. Proteins 17:87–92
Delaglio F, Grzesiek S, Vuister GW, Zhu G, Pfeifer J, Bax A (1995) NMRpipe—a multidimensional spectral processing system based on Unix pipes. J Biomol NMR 6:277–293
Dobson CM, Lian L-Y, Redfield C, Topping KD (1986) Measurement of hydrogen exchange rates using 2D NMR spectroscopy. J Magn Reson 69:201–209
Englander SW, Kallenbach NR (1983) Hydrogen exchange and structural dynamics of proteins and nucleic-acids. Q Rev Biophys 16:521–655
Fitzkee NC, Torchia DA, Bax A (2011) Measuring rapid hydrogen exchange in the homodimeric 36 kDa HIV-1 integrase catalytic core domain. Protein Sci 20:500–512
Geen H, Freeman R (1991) Band-selective radiofrequency pulses. J Magn Reson 93:93–141
Gemmecker G, Jahnke W, Kessler H (1993) Measurement of fast proton-exchange rates in isotopically labeled compounds. J Am Chem Soc 115:11620–11621
Grzesiek S, Bax A (1993) Measurement of amide proton exchange rates and NOEs with water in 13C/15 N-enriched calcineurin B. J Biomol NMR 3:627–638
Hwang TL, Mori S, Shaka AJ, van Zijl PCM (1997) Application of phase-modulated CLEAN chemical EXchange spectroscopy (CLEANEX-PM) to detect water-protein proton exchange and intermolecular NOEs. J Am Chem Soc 119:6203–6204
Hwang TL, van Zijl PCM, Mori S (1998) Accurate quantitation of water-amide proton exchange rates using the Phase-Modulated CLEAN chemical EXchange (CLEANEX-PM) approach with a Fast-HSQC (FHSQC) detection scheme. J Biomol NMR 11:221–226
Jeener J, Meier BH, Bachmann P, Emst RR (1979) Investigation of exchange processes by two-dimensional NMR spectroscopy. J Chem Phys 71:4546–4553
Kay LE, Keifer P, Saarinen T (1992) Pure absorption gradient enhanced heteronuclear single quantum correlation spectroscopy with improved sensitivity. J Am Chem Soc 114:10663–10665
Liepinsh E, Otting G (1995) Selective excitation of intense solvent signals in the presence of radiation damping. J Biomol NMR 5:420–426
Lim J, Rong K, Murugan E, Ho CL, Liang Z, Yang D (2011) Solution structures of the acyl carrier protein domain from the highly reducing Type I iterative polyketide synthase CalE8. Plos One 6:e20549
Lipari G, Szabo A (1982) Model-free approach to the interpretation of nuclear magnetic relaxation in macromolecules: 1. Theory and range of validity. J Am Chem Soc 104:4546–4559
Mao XA, Ye C (1997) Understanding radiation damping in a simple way. Concepts Magn Reson 9:173–187
Miao XJ, Jin-Hong Chen JH, Mao XA (1999) Selective excitation by radiation damping field for a coupled nuclear spin system. Chem Phys Lett 304:45–50
Mori S, Johnson MO, Berg JM, Vanzijl PCM (1994) Water exchange filter (wex filter) for nuclearmagnetic-resonance studies of macromolecules. J Am Chem Soc 116:11982–11984
Mori S, Berg JM, van Zijl PCM (1996a) Separation of intramolecular NOE and exchange peaks in water exchange spectroscopy using spin-echo filters. J Biomol NMR 7:77–82
Mori S, Abeygunawardana C, van Zijl PCM, Berg JM (1996b) Water exchange filter with improved sensitivity (WEX II) to study solvent-exchangeable protons. Application to the consensus zinc finger peptide CP-1. J Magn Reson B 110:96–101
Mori S, Abeygunawardana C, Berg JM, Peter CM, van Zijl PCM (1997) NMR study of rapidly exchanging backbone amide protons in staphylococcal nuclease and the correlation with structural and dynamic properties. J Am Chem Soc 119:6844–6852
Otting G, Liepinsh E, Wüthrich K (1991) Protein hydration in aqueous solution. Science 254:974–980
Parris KD, Lin L, Tam A, Mathew R, Hixon J, Stahl M, Fritz CC, Seehra J, Somers WS (2000) Crystal structures of substrate binding to Bacillus subtilis holo-(acyl carrier protein) synthase reveal a novel trimeric arrangement of molecules resulting in three active sites. Structure 8:883–895
Paterson Y, Englander SW, Roder H (1990) An antibody binding site on cytochrome c defined by hydrogen exchange and two-dimensional NMR. Science 249(4970):755–759
Spera S, Ikura M, Bax A (1991) Measurement of the exchange rates of rapidly exchanging amide protons. Application to the study of calmodulin and its complex with a myosin light chain kinase fragment. J Biomol NMR 1:155–166
Vijay-Kumar S, Charles E, Bugg CE, Cook WJ (1987) Structure of ubiquitin refined at 1.8 Å resolution. J Mol Biol 194:531–544
Wagner G, Wüthrich K (1982) Amide proton exchange and surface conformation of the basic pancreatic trypsin inhibitor in solution: studies with two-dimensional nuclear magnetic resonance. J Mol Biol 160:343–361
Acknowledgments
This work was supported by a grant from the ministry of education, Singapore (R154000453112). The authors thank Lewis E. Kay for his seminal contributions to the NMR community. DY is immensely grateful to Lewis for his patient guidance in NMR experiment developments and data analyses when DY was working in his laboratory. DY is indebted to Lewis for his continuous support, encouragement and inspiration.
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Fan, JS., Lim, J., Yu, B. et al. Measurement of amide hydrogen exchange rates with the use of radiation damping. J Biomol NMR 51, 151 (2011). https://doi.org/10.1007/s10858-011-9549-6
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DOI: https://doi.org/10.1007/s10858-011-9549-6