title: Simultaneous determination of ACE activity with 2 substrates provides information on the status of somatic ACE and allows detection of inhibitors in human blood.
abstract: Angiotensin I-converting enzyme (ACE), a key enzyme in cardiovascular pathophysiology, consists of 2 homologous domains, each bearing a Zn-dependent active site. The ratio of the rates of hydrolysis of 2 synthetic substrates, Z-Phe-His-Leu (ZPHL) and Hip-His-Leu (HHL), is characteristic for each type of ACE: somatic 2-domain 1, N-domain 4.5, and C-domain 0.7 (Williams et al, 1996). We hypothesized that inactivation or selective inhibition of the C-domain within the somatic ACE should increase the ratio from 1 toward higher values, whereas inactivation or inhibition of the N-domain should decrease the ratio to lower values. Temperatures in the 40-60 degrees C range cause preferential inactivation of the C-domain in somatic ACE and an increase in the ZPHL/HHL ratio. Determination of the ZPHL/HHL ratio in freshly 100-fold concentrated urine ruled out the existence of the N-domain fragment in human urine, which appears only as a result of long storage. Inhibition of ACE by common inhibitors also increases the ZPHL/HHL ratio for 2-domain ACE, thus providing a sensitive method for the detection of ACE inhibitors in biological fluids. Therefore, simultaneous measurements of ACE activity with 2 substrates (ZPHL and HHL) and calculation of their ratio allow us to monitor the status of the ACE molecule and detect ACE inhibitors (endogenous and exogenous) in human blood and other biological fluids. This method should find wide application in monitoring clinical trials with ACE inhibitors and in the development of the approach for personalized medicine by these effective drugs.
authors.affiliation: Department of Anesthesiology, University of Illinois at Chicago, Chicago, Illinois 60612, USA. danilov@uic.edu
authors.forename: Sergei M
authors.initials: SM
authors.lastname: Danilov
authors.forename: Irina V
authors.initials: IV
authors.lastname: Balyasnikova
authors.forename: Ronald F
authors.initials: RF
authors.lastname: Albrecht
authors.forename: Olga A
authors.initials: OA
authors.lastname: Kost
mesh_terms: D000806:Angiotensin-Converting Enzyme Inhibitors / Q000097:blood* / Q000652:urine; D015153:Blotting, Western; D006801:Humans; D007700:Kinetics; D009842:Oligopeptides / Q000378:metabolism; D017434:Protein Structure, Tertiary; D013379:Substrate Specificity
chemical_list: D000806:Angiotensin-Converting Enzyme Inhibitors; C477729:N-carbobenzoxy-phenylalanyl-histidyl-leucine; D009842:Oligopeptides; C010980:hippuryl-histidyl-leucine