A novel metalloproteinase present in freshly isolated rat glomeruli.
We have utilized [3H] gelatin to document high activity of a metalloproteinase present in freshly isolated rat glomeruli. [3H] gelatin degradation by glomeruli was markedly inhibited by EDTA (10 mM: -89 +/- 2.3%) and o-phenanthroline (2 mM: -72 +/- 0.1%), inhibitors of metalloproteinases. No significant inhibition of [3H]gelatin degradation was observed with inhibitors of serine or cysteine proteinases. Most (greater than 80%) of the glomerular metalloproteinase (GLOMP) activity was associated with the pellet after centrifugation of sonicated glomeruli at 100,000 g for 90 min. The pH optimum for gelatin degradation by sonicated glomeruli was approximately pH 8.5. Sodium dodecyl sulfate substrate (gelatin)-polyacrylamide gel electrophoresis revealed a single major band of EDTA-inhibitable gelatin-degrading activity with a molecular mass of approximately 116-125 kDa. The GLOMP activity was not inhibited by tissue inhibitors of metalloproteinases, did not appear to be latent, and was not activated by organomercurial activators of several latent metalloproteinases. GLOMP activity was increased 3.4-fold after incubation with trypsin (20 micrograms/ml, 25 min, 22 degrees C). These data indicate that GLOMP is distinct from the previously described matrix metalloproteinases, as well as other metalloproteinases present in the kidney, including the gelatinase secreted by cultured mesangial cells, Meprin, and endopeptidase 24.11 (enkephalinase, EC 3.4.24.11).
Le
Q
Q
Department of Biochemistry, Tulane Medical School, New Orleans, Louisiana 70112.
Shah
S
S
Nguyen
H
H
Cortez
S
S
Baricos
W
W
D000818:Animals; D002458:Cell Fractionation; D004229:Dithiothreitol / Q000494:pharmacology; D004492:Edetic Acid / Q000494:pharmacology; D004591:Electrophoresis, Polyacrylamide Gel; D004789:Enzyme Activation / Q000187:drug effects; D005780:Gelatin / Q000378:metabolism; D006023:Glycoproteins / Q000494:pharmacology; D006863:Hydrogen-Ion Concentration; D007668:Kidney / Q000201:enzymology; D007678:Kidney Glomerulus / Q000201:enzymology*; D008666:Metalloendopeptidases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D008970:Molecular Weight; D010618:Phenanthrolines / Q000494:pharmacology; D051381:Rats; D019714:Tissue Inhibitor of Metalloproteinases; D014357:Trypsin / Q000494:pharmacology
D006023:Glycoproteins; D010618:Phenanthrolines; D019714:Tissue Inhibitor of Metalloproteinases; D005780:Gelatin; D004492:Edetic Acid; D014357:Trypsin; D008666:Metalloendopeptidases; D004229:Dithiothreitol; C025205:1,10-phenanthroline