Degradation of native type IV procollagen by human neutrophil elastase. Implications for leukocyte-mediated degradation of basement membranes.
Leukocyte-derived proteases may contribute to the destruction of basement membranes during inflammation. We have, therefore, examined the degradation of human type IV procollagen (PC) by purified human neutrophil elastase (HLE). Native [14C]proline-labeled type IV PC was isolated from cultures of human HT-1080 cells and incubated with HLE for various times at 25 or 37 degrees C. Cleavage products were resolved by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and identified by CNBr peptide mapping. Incubation of type IV PC with HLE (less than 1:10 HLE:type IV weight ratio) resulted in cleavage of the pro alpha 1 (IV) and pro alpha 2 chains (Mr 180,000 and 175,000) to discrete components of Mr greater than 140,000. Peptide mapping indicated that the carboxy-terminal collagenase-resistant domains of both chains were rapidly and preferentially degraded. Longer incubations or incubations at higher enzyme:substrate ratios resulted in extensive and asymmetric internal cleavage with the generation of fragments similar in size distribution to the major pepsin-resistant fragments of type IV collagen. Our findings indicate that soluble, native human type IV PC is a substrate for HLE and is preferentially cleaved within the globular carboxy-terminal domains of the pro alpha 1 and pro alpha 2 chains. We suggest that even limited cleavage of type IV PC by HLE may disrupt intermolecular carboxy-terminal interactions believed to be important for basement membrane assembly and for maintaining basement membrane structure in vivo.
Pipoly
D J
DJ
Department of Pathology, Jewish Hospital of St. Louis, Washington University Medical Center, Missouri 63110.
Crouch
E C
EC
D001485:Basement Membrane / Q000378:metabolism*; D002460:Cell Line; D006801:Humans; D007249:Inflammation; D007700:Kinetics; D009504:Neutrophils / Q000201:enzymology* / Q000502:physiology; D010084:Oxidation-Reduction; D010196:Pancreatic Elastase / Q000097:blood*; D011347:Procollagen / Q000378:metabolism*
D011347:Procollagen; D010196:Pancreatic Elastase