Partial purification and characterization of a rat kidney neutral endopeptidase that hydrolyzes succinyl trialanine-4-nitroanilide.
The activity for the hydrolysis of succinyl trialanine-4-nitroanilide was higher in kidney homogenates of female rats and mice than in those of male rats and mice. An enzyme hydrolyzing the above substrate was extracted from female rat kidney homogenate and partially purified by means of gel filtration on Sepharose 4B, anion-exchange chromatography on DEAE-Sepharose CL-6B and affinity chromatography on carbobenzoxy-L-Ala-L-Ala-D-Ala-polylysine-agarose. The purified enzyme cleaved the bond between succinyl dialanine and alanine-4-nitroanilide of the substrate and showed a Km value of 3.3 mM at the optimal pH of 7.5. The activity was increased by Ca2+ and Mg2+, but inhibited by EDTA. With oxidized insulin B chain as a substrate, the enzyme cleaved the carbonyl bonds of Ala-14, Tyr-16 and Gly-23 efficiently, and those of His-5 and His-10 less efficiently.
D000595:Amino Acid Sequence; D000818:Animals; D003921:Diabetes Mellitus, Experimental / Q000201:enzymology; D010450:Endopeptidases / Q000302:isolation & purification* / Q000378:metabolism; D007328:Insulin / Q000378:metabolism; D007668:Kidney / Q000201:enzymology*; D008099:Liver / Q000201:enzymology; D051379:Mice; D008810:Mice, Inbred C57BL; D008813:Mice, Inbred ICR; D008817:Mice, Mutant Strains; D015260:Neprilysin; D010179:Pancreas / Q000201:enzymology; D051381:Rats; D011919:Rats, Inbred Strains; D013045:Species Specificity; D013379:Substrate Specificity
D007328:Insulin; D010450:Endopeptidases; D015260:Neprilysin