CaltechAUTHORS
  A Caltech Library Service

Nitrogen-15 nuclear magnetic resonance spectroscopy. The state of histidine in the catalytic triad of .alpha.-lytic protease. Implications for the charge-relay mechanism of peptide-bond cleavage by serine proteases

Bachovchin, William W. and Roberts, John D. (1978) Nitrogen-15 nuclear magnetic resonance spectroscopy. The state of histidine in the catalytic triad of .alpha.-lytic protease. Implications for the charge-relay mechanism of peptide-bond cleavage by serine proteases. Journal of the American Chemical Society, 100 (26). pp. 8041-8047. ISSN 0002-7863. http://resolver.caltech.edu/CaltechAUTHORS:20151007-093103668

Full text is not posted in this repository. Consult Related URLs below.

Use this Persistent URL to link to this item: http://resolver.caltech.edu/CaltechAUTHORS:20151007-093103668

Abstract

Histidine, enriched in ^(15)N in the imidazole ring, has been incorporated into the “catalytic triad“ of serinyl. histidyl, and aspartyl residues of α-lytic protease, using a histidine auxotroph of myxobacter 495. The pK_a of this histidyl residue is 7.0 ± 0.1 at 26 ºC, as determined by the changes of its ^(15)N chemical shifts in nuclear magnetic resonance spectra. This finding is contrary to previously published reports that histidyl residues at the active sites of serine proteases are likely to be abnormally weak bases, while the “buried” aspartyl residues of the catalytic triads are likely to be abnormally weak acids and thus offers no support for the current formulation of the charge-relay mechanism of action of serine proteases. The ^(15)N chemical shifts further demonstrate that, at catalytically active pH values, the tautomer with hydrogen on N3 (the π nitrogen) of the imidazole ring predominates. This is an unusual tautomeric state both for simple 4-substituted imidazole derivatives and for histidyl residues in proteins, and its predominance in ɑ-lytic protease can be reasonably attributed to a hydrogen-bonded interaction between NH at the 3 position and the adjacent “buried” carboxylate group of aspartic acid. The implication of the results for the respective roles of the aspartyl and histidyl residues in serine protease catalysis is discussed in terms of the energy requirements of the individual steps for the formation and breakdown of the serine ester intermediate.


Item Type:Article
Related URLs:
URLURL TypeDescription
http://dx.doi.org/10.1021/ja00494a001DOIArticle
http://pubs.acs.org/doi/abs/10.1021/ja00494a001PublisherArticle
Additional Information:© 1978 American Chemical Society. Received May 26, 1978. Supported by the Public Health Service, Research Grant No. GM-11072, from the Division of General Medical Sciences, and by the National Science Foundation. We are greatly indebted to Professor John H. Richards and Dr. Michael W. Hunkapiller for their unfailing interest and assistance throughout the course of this research, but we should emphasize that they are in no way responsible for its deficiencies either in the experiments or our interpretation of the results. We also appreciate the myxobacter 495 culture supplied by Dr. Hunkapiller, from which the auxotroph was derived.
Funders:
Funding AgencyGrant Number
U.S. Public Health Service (USPHS)GM-11072
NSFUNSPECIFIED
Other Numbering System:
Other Numbering System NameOther Numbering System ID
Caltech Gates and Crellin Laboratories of Chemistry5768
Record Number:CaltechAUTHORS:20151007-093103668
Persistent URL:http://resolver.caltech.edu/CaltechAUTHORS:20151007-093103668
Official Citation:Nitrogen-15 nuclear magnetic resonance spectroscopy. The state of histidine in the catalytic triad of .alpha.-lytic protease. Implications for the charge-relay mechanism of peptide-bond cleavage by serine proteases William W. Bachovchin and John D. Roberts Journal of the American Chemical Society 1978 100 (26), 8041-8047 DOI: 10.1021/ja00494a001
Usage Policy:No commercial reproduction, distribution, display or performance rights in this work are provided.
ID Code:60864
Collection:CaltechAUTHORS
Deposited By: Ruth Sustaita
Deposited On:07 Oct 2015 22:37
Last Modified:07 Oct 2015 22:37

Repository Staff Only: item control page