Comparative sequence specificities of human 72- and 92-kDa gelatinases (type IV collagenases) and PUMP (matrilysin).
The sequence specificities of human 72-kDa fibroblast gelatinase (type IV collagenase), human 92-kDa neutrophil gelatinase (type IV collagenase), and putative metalloproteinase (PUMP or matrilysin) have been examined by measuring the rate of hydrolysis of over 50 synthetic oligopeptides covering the P4 through P4' subsites of the substrate. The peptides investigated in this paper were those employed in our previous study which systematically examined the sequence specificity of human fibroblast and neutrophil collagenases [Netzel-Arnett et al. (1991) J. Biol. Chem. 266, 6747]. The initial rate of hydrolysis of the P1-P1' bond of each peptide has been measured under first-order conditions ([S0] << KM), and kcat/KM values have been calculated from the initial rates. The specificities of these five metalloproteinases are similar, but distinct, with the largest differences occurring at subsites P1, P1', and P3'. The specificities of the two gelatinases are the most similar to each other. They tolerate only small amino acids such as Gly and Ala in subsite P1. In contrast, larger residues such as Met, Pro, Gln, and Glu are also accommodated well by PUMP. All five enzymes prefer hydrophobic, aliphatic residues in subsite P1'. PUMP exhibits a stronger preference for Leu in this subsite than is shown by the other enzymes. The P3' subsite specificities of the gelatinases and collagenases are very similar but different from those of PUMP which particularly prefers Met in this position. The specificity data from this study allow the design of optimized substrates and selective inhibitors for these metalloproteinases.
Department of Chemistry, Florida State University, Tallahassee 32306.
S
S
Netzel-Arnett
Q X
QX
Sang
W G
WG
Moore
M
M
Navre
H
H
Birkedal-Hansen
H E
HE
Van Wart
D053674:Aggrecans; D000595:Amino Acid Sequence; D011508:Chondroitin Sulfate Proteoglycans / Q000378:metabolism; D003094:Collagen / Q000378:metabolism; D010450:Endopeptidases / Q000378:metabolism*; D016326:Extracellular Matrix Proteins; D018093:Gelatinases; D006801:Humans; D007700:Kinetics; D037181:Lectins, C-Type; D020783:Matrix Metalloproteinase 7; D008666:Metalloendopeptidases / Q000378:metabolism*; D008969:Molecular Sequence Data; D008970:Molecular Weight; D009842:Oligopeptides / Q000378:metabolism*; D011509:Proteoglycans / Q000378:metabolism; D013329:Structure-Activity Relationship; D013379:Substrate Specificity
D053674:Aggrecans; D011508:Chondroitin Sulfate Proteoglycans; D016326:Extracellular Matrix Proteins; D037181:Lectins, C-Type; D009842:Oligopeptides; D011509:Proteoglycans; D003094:Collagen; D010450:Endopeptidases; D018093:Gelatinases; D008666:Metalloendopeptidases; D020783:Matrix Metalloproteinase 7