A sensitive and rapid method for assaying the activity of type III procollagen amino-terminal proteinase.
A rapid assay procedure was developed for measuring the rate of cleavage of the amino-terminal propeptide of type III procollagen. The method was based on the sequential precipitation of type III collagen and uncleaved pN-collagen by 30% ammonium sulfate, while the free amino-terminal propeptide remained in solution and could be further precipitated by 60% ammonium sulfate. Consistently better results were obtained than with the earlier method in which absolute ethanol was used as the precipitant, and selective precipitation was confirmed by polyacrylamide gel electrophoresis of the pellets. The high sensitivity of this method facilitates relatively rapid assays even from small amounts of cultured cells.
D000596:Amino Acids / Q000032:analysis; D000645:Ammonium Sulfate; D000818:Animals; D001011:Aorta / Q000201:enzymology; D002417:Cattle; D002478:Cells, Cultured; D011232:Chemical Precipitation; D003094:Collagen / Q000032:analysis; D010450:Endopeptidases / Q000032:analysis* / Q000302:isolation & purification; D005347:Fibroblasts / Q000201:enzymology; D006801:Humans; D009131:Muscle, Smooth, Vascular / Q000201:enzymology; D010446:Peptide Fragments / Q000032:analysis; D011347:Procollagen / Q000032:analysis; D011348:Procollagen N-Endopeptidase
D000596:Amino Acids; D010446:Peptide Fragments; D011347:Procollagen; C041876:procollagen Type III-N-terminal peptide; D003094:Collagen; D010450:Endopeptidases; D011348:Procollagen N-Endopeptidase; D000645:Ammonium Sulfate