13C- and 1H-NMR studies of oxyanion and tetrahedral intermediate stabilization by the serine proteinases: optimizing inhibitor warhead specificity and potency by studying the inhibition of the serine proteinases by peptide-derived chloromethane and glyoxal inhibitors

Biochem Soc Trans. 2007 Jun;35(Pt 3):566-70. doi: 10.1042/BST0350566.

Abstract

Catalysis by the serine proteinases proceeds via a tetrahedral intermediate whose oxyanion is stabilized by hydrogen-bonding in the oxyanion hole. There have been extensive (13)C-NMR studies of oxyanion and tetrahedral intermediate stabilization in trypsin, subtilisin and chymotrypsin using substrate-derived chloromethane inhibitors. One of the limitations of these inhibitors is that they irreversibly alkylate the active-site histidine residue which results in the oxyanion not being in the optimal position in the oxyanion hole. Substrate-derived glyoxal inhibitors are reversible inhibitors which, if they form tetrahedral adducts in the same way as substrates form tetrahedral intermediates, will overcome this limitation. Therefore we have synthesized (13)C-enriched substrate-derived glyoxal inhibitors which have allowed us to use (13)C-NMR and (1)H-NMR to determine how they interact with proteinases. It is hoped that these studies will help in the design of specific and highly potent warheads for serine proteinase inhibitors.

Publication types

  • Research Support, Non-U.S. Gov't
  • Review

MeSH terms

  • Amino Acid Sequence
  • Chymotrypsin / antagonists & inhibitors
  • Chymotrypsin / chemistry
  • Chymotrypsin / metabolism
  • Enzyme Stability
  • Glyoxal / pharmacology
  • Hydrogen Bonding
  • Hydrogen-Ion Concentration
  • In Vitro Techniques
  • Methyl Chloride / pharmacology
  • Nuclear Magnetic Resonance, Biomolecular
  • Oligopeptides / chemistry
  • Oligopeptides / metabolism
  • Protons
  • Serine Endopeptidases / chemistry*
  • Serine Endopeptidases / metabolism*
  • Serine Proteinase Inhibitors / pharmacology
  • Subtilisin / metabolism

Substances

  • Oligopeptides
  • Protons
  • Serine Proteinase Inhibitors
  • Glyoxal
  • Methyl Chloride
  • Serine Endopeptidases
  • Chymotrypsin
  • Subtilisin