NMR study of rapidly exchanging backbone amide protons in staphylococcal nuclease and the correlation with structural arid dynamic properties

Susumu Mori, Chitrananda Abeygunawardana, Jeremy M. Berg, Peter C.M. Van Zijl

Research output: Contribution to journalArticlepeer-review

Abstract

Exchange rates of rapidly exchanging (>1.0 s-1) backbone amide protons with solvent water in staphylococcal nuclease (SN) were measured at pH 6.03- 7.03 with a 2D heteronuclear water exchange filter sequence (WEX II-FHSQC). Comparison of the exchange data with crystal structure data reveals the following: (1) Non-hydrogen-bonding and exposed residues have protection factors (predicted exchange rate in random coil/measured exchange rate) of less than 15 for non-hydrophobic residues and 10 or higher for hydrophobic residues. Among non-hydrophobic residues, Gly tends to have a higher protection factor (10-15), whereas other residues are below 10. (2) Protection factors for buried and non-hydrogen-bonded protons vary over a wide range (6-104). Low protection factors (<25) may indicate fluctuations in structure resulting in substantial solvent exposure. (3) Some hydrogen- bonded and buried protons show rapid exchange, and the protection factors are 25-400. This indicates kinetically labile hydrogen bonds and solvent exposure by structural fluctuations. On the other hand, many protons in this category exchange slower than the detection limit (<1.0 s-1) and are mainly observed in α helices and β sheets, or hydrophobic residues. Although a good correlation between NMR exchange measurements and X-ray structural properties was observed, discrepancies were also found for several residues, namely 80Gln, 82Thr, and 138Asn. The measured pH dependence revealed unusual behavior for 77Asn, possibly due to a tightly bound water molecule. Our data indicate that it is possible to use the exchange rates of rapidly exchanging protons as a probe for studying hydrogen bonding, solvent accessibility, or regional kinetic lability of protein structures in solution at physiological pH.

Original languageEnglish (US)
Pages (from-to)6844-6852
Number of pages9
JournalJournal of the American Chemical Society
Volume119
Issue number29
DOIs
StatePublished - Jul 23 1997

ASJC Scopus subject areas

  • Catalysis
  • Chemistry(all)
  • Biochemistry
  • Colloid and Surface Chemistry

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