Characterization of 58-kilodalton human neutrophil collagenase: comparison with human fibroblast collagenase.
A series of experiments has been carried out to characterize 58-kDa human neutrophil collagenase (HNC) and compare it with human fibroblast collagenase (HFC). N-Terminal sequencing of latent and spontaneously activated HNC shows that it is a distinct collagenase that is homologous to HFC and other members of the matrix metalloproteinase gene family. Activation occurs autolytically by hydrolysis of an M-L bond at a locus homologous to the Q80-F81-V82-L83 autolytic activation site of HFC. This releases a 16-residue propeptide believed to contain the "cysteine switch" residue required for latency. Polyclonal antibody raised against HNC cross-reacts with HFC but with none of the other major human matrix metalloproteinases examined. Treatment of HNC with endoglycosidase F or N-glycosidase F indicates that it is glycosylated at multiple sites. The deglycosylated latent and spontaneously activated enzymes have molecular weights of approximately 44K and 42K, respectively. Differences in the carbohydrate processing of HFC and HNC may determine why HFC is a secreted protein while HNC is stored in intracellular granules. The kinetic parameters kcat and KM for the hydrolysis of the interstitial collagen types I, II, and III in solution by both collagenases have been determined. The strong preferences of HNC for type I collagen and of HFC for type III collagen found in earlier studies have been confirmed. The preference of HNC for type I over type III collagen is almost abolished when fibrillar collagens are used as substrates, but the preference for HFC for type III over type I collagen is only partially decreased.(ABSTRACT TRUNCATED AT 250 WORDS)
Mallya
S K
SK
Department of Chemistry, Florida State University, Tallahassee 32306.
Mookhtiar
K A
KA
Gao
Y
Y
Brew
K
K
Dioszegi
M
M
Birkedal-Hansen
H
H
Van Wart
H E
HE
D000595:Amino Acid Sequence; D015153:Blotting, Western; D003429:Cross Reactions; D005347:Fibroblasts / Q000201:enzymology*; D006023:Glycoproteins; D006801:Humans; D003012:Microbial Collagenase / Q000737:chemistry / Q000378:metabolism / Q000502:physiology*; D008969:Molecular Sequence Data; D008970:Molecular Weight; D009504:Neutrophils / Q000201:enzymology*; D011499:Protein Processing, Post-Translational; D013379:Substrate Specificity
D006023:Glycoproteins; D003012:Microbial Collagenase