Apparent kinetics of angiotensin converting enzyme: hydrolysis of [3H]benzoyl-phenylalanyl-alanyl-proline in the isolated perfused lung.
Isolated perfused rabbit lungs were used to study the hydrolysis of [3H]benzoyl-phenylalanyl-alanyl-proline [( 3H]BPAP), a synthetic substrate for angiotensin converting enzyme (ACE). Lungs were perfused, at constant flow rates, with physiologic medium containing added BPAP and the concentration of its metabolite, [3H]benzoyl-phenylalaline, was measured in lung effluent. Hydrolysis of BPAP (4.2 microM) was 64.1 +/- 3.3% at 37 degrees C and was significantly decreased (P less than .01) to 10.1 +/- 8.7% by the addition of the ACE inhibitor, MK422 (10(-6) M). Disappearance (i.e., hydrolysis) of immunoreactive angiotensin I was also inhibited by MK422. Hydrolysis of BPAP was saturable and calculated apparent kinetic constants were Km = 13 microM and Vmax = 50 nmol/sec/lung. When the perfusion medium temperature was 10 degrees C, apparent Km was unchanged, whereas Vmax was significantly (P less than .05) decreased. At BPAP concentrations sufficient to depress metabolism to less than 20%, perfusion pressure was unchanged. Hydrolysis of BPAP under first-order conditions was independent of flow over the range 10 to 50 ml/min. However, increase in flow rate to 100 ml/min/lung was associated with decreased BPAP metabolism. These data indicate that BPAP is a substrate for pulmonary ACE in vitro and substantiate its use in intact animals because: 1) it is without physiologic effect in high doses; and 2) calculated apparent kinetics in isolated lungs under these conditions of steady-state concentrations were similar to those obtained from earlier studies that utilized bolus injection techniques in intact animals.
Ashton
J H
JH
Pitt
B R
BR
Gillis
C N
CN
D000803:Angiotensin I / Q000378:metabolism; D000806:Angiotensin-Converting Enzyme Inhibitors; D000818:Animals; D004151:Dipeptides / Q000494:pharmacology; D015773:Enalaprilat; D006868:Hydrolysis; D007700:Kinetics; D008168:Lung / Q000201:enzymology*; D008297:Male; D008433:Mathematics; D009842:Oligopeptides / Q000378:metabolism*; D007703:Peptidyl-Dipeptidase A / Q000378:metabolism*; D010477:Perfusion; D011817:Rabbits; D012701:Serotonin / Q000378:metabolism; D013696:Temperature
D000806:Angiotensin-Converting Enzyme Inhibitors; D004151:Dipeptides; D009842:Oligopeptides; D012701:Serotonin; C029689:benzoylphenylalanyl-alanyl-proline; D000803:Angiotensin I; D007703:Peptidyl-Dipeptidase A; D015773:Enalaprilat