Electronic spectroscopy of cobalt angiotensin converting enzyme and its inhibitor complexes.
Zinc, the catalytically essential metal of angiotensin converting enzyme (ACE), has been replaced by cobalt(II) to give an active, chromophoric enzyme that is spectroscopically responsive to inhibitor binding. Visible absorption spectroscopy and magnetic circular dichroic spectropolarimetry have been used to characterize the catalytic metal binding site in both the cobalt enzyme and in several enzyme-inhibitor complexes. The visible absorption spectrum of cobalt ACE exhibits a single broad maximum (525 nm) of relatively low absorptivity (epsilon = 75 M-1 cm-1). In contrast, the spectra of enzyme-inhibitor complexes display more clearly defined maxima at longer wavelengths (525-637 nm) and of markedly higher absorptivities (130-560 M-1 cm-1). The large spectral response indicates that changes in the cobalt ion coordination sphere occur on inhibitor binding. Magnetic circular dichroic spectropolarimetry has shown that the metal coordination geometry in the inhibitor complexes is tetrahedral and of higher symmetry than in cobalt ACE alone. The presence of sulfur----cobalt charge-transfer bands in both the visible absorption and magnetic circular dichroic spectra of the cobalt ACE-Captopril complex confirm direct ligation of the thiol group of the inhibitor to the active-site metal.
Bicknell
R
R
Center for Biochemical and Biophysical Sciences and Medicine, Harvard Medical School, Boston, Massachusetts.
Holmquist
B
B
Lee
F S
FS
Martin
M T
MT
Riordan
J F
JF
D000806:Angiotensin-Converting Enzyme Inhibitors; D000818:Animals; D002216:Captopril / Q000494:pharmacology; D003035:Cobalt / Q000494:pharmacology*; D004656:Enalapril / Q000031:analogs & derivatives / Q000494:pharmacology; D015773:Enalaprilat; D004789:Enzyme Activation / Q000187:drug effects; D007700:Kinetics; D008168:Lung / Q000201:enzymology; D007703:Peptidyl-Dipeptidase A; D011485:Protein Binding; D011817:Rabbits; D013053:Spectrophotometry / Q000379:methods
D000806:Angiotensin-Converting Enzyme Inhibitors; D003035:Cobalt; D004656:Enalapril; D002216:Captopril; D007703:Peptidyl-Dipeptidase A; D015773:Enalaprilat