Thermal denaturation of rat pulmonary and testicular angiotensin-converting enzyme isozymes. Effects of chelators and CoCl2.
In an attempt to assess the biochemical consequences resulting from structural differences between rat pulmonary and testicular angiotensin-converting enzyme, the thermal stability of crude and purified preparations of each enzyme was compared. Structural heterology was verified by molecular weight determinations and by peptide mapping after limited proteolysis with Staphylococcus V8 proteinase. Thermal stability was monitored by changes in catalytic activity following incubations at 55 degrees C in the presence of chelators and CoCl2. Purified pulmonary angiotensin-converting enzyme was more sensitive to inhibition by the chelators EDTA and 1,10-phenanthroline and by the site-directed inhibitor captopril than was the testicular isozyme. Although the pulmonary holoenzyme was unaffected by cobalt, the testicular holoenzyme was inhibited by cobalt in a concentration-dependent manner. Crude pulmonary angiotensin-converting enzyme was significantly more resistant to thermal denaturation than its crude testicular counterpart. The differences in the thermal lability of each isozyme were still present in purified preparations, although the purified enzymes appeared to be more thermally stable than their crude counterparts. Both chelators and cobalt markedly potentiated the thermal denaturation of each isozyme. These data suggest that the structural heterology of the pulmonary and testicular isozymes may affect the interaction of zinc with the respective enzymes and that zinc may contribute to the structural integrity and thermal stability of angiotensin-converting enzyme in each tissue.
Velletri
P A
PA
Billingsley
M L
ML
Lovenberg
W
W
D000806:Angiotensin-Converting Enzyme Inhibitors; D000818:Animals; D002614:Chelating Agents / Q000494:pharmacology; D003035:Cobalt / Q000494:pharmacology; D006358:Hot Temperature; D066298:In Vitro Techniques; D007527:Isoenzymes / Q000037:antagonists & inhibitors*; D008168:Lung / Q000201:enzymology*; D008297:Male; D009928:Organ Specificity; D011489:Protein Denaturation; D051381:Rats; D011919:Rats, Inbred Strains; D013737:Testis / Q000201:enzymology*
D000806:Angiotensin-Converting Enzyme Inhibitors; D002614:Chelating Agents; D007527:Isoenzymes; D003035:Cobalt; C018021:cobaltous chloride