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A Fluorescence‐Based Array Screen for Transglutaminase Substrates
Author(s) -
Malešević Miroslav,
Migge Andreas,
Hertel Thomas C.,
Pietzsch Markus
Publication year - 2015
Publication title -
chembiochem
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.05
H-Index - 126
eISSN - 1439-7633
pISSN - 1439-4227
DOI - 10.1002/cbic.201402709
Subject(s) - tripeptide , tissue transglutaminase , lysine , chemistry , biochemistry , glutamine , amino acid , peptide , substrate (aquarium) , enzyme , peptide bond , arginine , biotinylation , amine gas treating , tyrosine , stereochemistry , biology , organic chemistry , ecology
Transglutaminases (EC 2.3.2.13) form an enzyme family that catalyzes the formation of isopeptide bonds between the γ‐carboxamide group of glutamine and the ε‐amine group of lysine residues of peptides and proteins. Other primary amines can be accepted in place of lysine. Because of their important physiological and pathophysiological functions, transglutaminases have been studied for 60 years. However, the substrate preferences of this enzyme class remain largely elusive. In this study, we used focused combinatorial libraries of 400 peptides to investigate the influence of the amino acids adjacent to the glutamine and lysine residues on the catalysis of isopeptide bond formation by microbial transglutaminase. Using the peptide microarray technology we found a strong positive influence of hydrophobic and basic amino acids, especially arginine, tyrosine, and leucine. Several tripeptide substrates were synthesized, and enzymatic kinetic parameters were determined both by microarray analysis and in solution.