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Inhibition of thermolysin by phosphonamidate transition-state analogues: measurement of 31P-15N bond lengths and chemical shifts in two enzyme-inhibitor complexes by solid-state nuclear magnetic resonance.

Publication ,  Journal Article
Copié, V; Kolbert, AC; Drewry, DH; Bartlett, PA; Oas, TG; Griffin, RG
Published in: Biochemistry
October 2, 1990

31P and 15N chemical shifts and 31P-15N bond lengths have been measured with solid-state NMR techniques in two inhibitors of thermolysin, carbobenzoxy-Glyp-L-Leu-L-Ala (ZGpLA) and carbobenzoxy-L-Phep-L-Leu-L-Ala (ZFpLA), both as free lithium salts and when bound to the enzyme. Binding of both inhibitors to thermolysin results in large changes in the 31P chemical shifts. These changes are more dramatic for the tighter binding inhibitor ZFpLA, where a approximately 20 ppm downfield movement of the 31P isotropic chemical shift (sigma iso) is observed. This shift is due to changes in the shift tensor elements sigma 11 and sigma 22, while sigma 33 remains essentially constant. We observed a similar pattern for ZGpLA, but only a approximately 5 ppm change occurs in sigma iso. The changes in the 15N chemical shifts for both inhibitors are small upon binding, amounting to downfield shifts of 2 and 4 ppm for ZGpLA and ZFpLA, respectively. This indicates that there are no changes in the protonation state of the 15N in either the ZFpLA- or the ZGpLA-thermolysin complex. NMR distance measurements yield a P-N bond length rP-N = 1.68 +/- 0.03 A for the tight binding inhibitor ZFpLA both in its free lithium salt form and in its thermolysin-ZFpLA complex, a distance that is much shorter than the 1.90-A distance reported by X-ray crystallography studies [Holden et al. (1987) Biochemistry 26, 8542-8553].(ABSTRACT TRUNCATED AT 250 WORDS)

Duke Scholars

Published In

Biochemistry

DOI

ISSN

0006-2960

Publication Date

October 2, 1990

Volume

29

Issue

39

Start / End Page

9176 / 9184

Location

United States

Related Subject Headings

  • X-Ray Diffraction
  • Thermolysin
  • Solutions
  • Phosphorus Isotopes
  • Oligopeptides
  • Nitrogen Isotopes
  • Magnetic Resonance Spectroscopy
  • Hydrogen Bonding
  • Biochemistry & Molecular Biology
  • Binding Sites
 

Citation

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Copié, V., Kolbert, A. C., Drewry, D. H., Bartlett, P. A., Oas, T. G., & Griffin, R. G. (1990). Inhibition of thermolysin by phosphonamidate transition-state analogues: measurement of 31P-15N bond lengths and chemical shifts in two enzyme-inhibitor complexes by solid-state nuclear magnetic resonance. Biochemistry, 29(39), 9176–9184. https://doi.org/10.1021/bi00491a011
Copié, V., A. C. Kolbert, D. H. Drewry, P. A. Bartlett, T. G. Oas, and R. G. Griffin. “Inhibition of thermolysin by phosphonamidate transition-state analogues: measurement of 31P-15N bond lengths and chemical shifts in two enzyme-inhibitor complexes by solid-state nuclear magnetic resonance.Biochemistry 29, no. 39 (October 2, 1990): 9176–84. https://doi.org/10.1021/bi00491a011.
Journal cover image

Published In

Biochemistry

DOI

ISSN

0006-2960

Publication Date

October 2, 1990

Volume

29

Issue

39

Start / End Page

9176 / 9184

Location

United States

Related Subject Headings

  • X-Ray Diffraction
  • Thermolysin
  • Solutions
  • Phosphorus Isotopes
  • Oligopeptides
  • Nitrogen Isotopes
  • Magnetic Resonance Spectroscopy
  • Hydrogen Bonding
  • Biochemistry & Molecular Biology
  • Binding Sites