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The 599‐611 loop in thimet oligopeptidase modulates substrate selection via hydrogen bonding
Author(s) -
Song Michelle M.,
Elmore Donald E.,
Sigman Jeffrey A.,
Bruce Lisa A.,
Pabon Amanda,
Glucksman Marc J.,
Wolfson Adele J.
Publication year - 2008
Publication title -
the faseb journal
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.709
H-Index - 277
eISSN - 1530-6860
pISSN - 0892-6638
DOI - 10.1096/fasebj.22.1_supplement.793.7
Subject(s) - chemistry , stereochemistry , active site , oligopeptidase , hydrogen bond , mutant , substrate (aquarium) , binding site , enzyme , biochemistry , biophysics , biology , gene , molecule , ecology , organic chemistry
Thimet oligopeptidase (TOP) is a metalloenzyme with a broad specificity for small bioactive neuropeptides. We have previously shown that flexibility in a glycine‐rich loop region comprised of residues 599‐611 modulates substrate selection and catalysis. Computer simulations reveal that mutations of Gly to Ala in the loop lead to decreased flexibility in this region of the enzyme proximal to the active site. Molecular dynamics simulations further indicate that the Gly to Ala mutations’ effect upon catalytic activity towards different substrates is due to a variation in the degree of hydrogen bonding between the substrates and catalytic residues, specifically Tyr609 and Tyr612, which are implicated in transition state stabilization. The mutants and wild type enzyme exhibit similar inhibitor binding affinity, and the substrate cleavage sites are the same for both mutant and wild‐type enzymes, indicating that neither binding affinity nor substrate alignment are affected by the mutations. We conclude that flexibility in the loop confers a selectivity to a variety of neuropeptide substrates by altering the level of hydrogen bonding to a given peptide during catalysis.