The Use of Paramagnetic and Fluorescent Quenching Amino Acid TOAC for Evaluating Angiotensin I-Converting Enzyme

摘要

Advantageously to other fluorescent quenching probes known so far, the TOAC (2,2,6,6-tetramethylpiperidine-l-oxyl-4-amino-4-carboxylic acid) spin label, introduced earlier by us in the chemistry of peptides [1,2] , can be inserted at any position of an enzyme substrate. Taking into account this characteristic, the present work examined the specificity of angiotensin Iconverting enzyme (ACE) that cleaves the vasoactive angiotensin I (AI, DRVYIHPFHL) to produce the angiotensin II and on the other hand, inactivates the hypotensor bradykinin (BK, RPPGFSPFR) [3,4]. For this evaluation, TOAC-attaching AI (at positions 0, 1, 3, 5, 8, 9 and 10) and BK (at positions 0, 3, 7 and 9) were synthesized and evaluated towards their properties as ACE substrates. Moreover, EPR and fluorescence investigations were also initiated with these labeled analogues aiming at further investigating the structure-function relationship and also the possibility of monitoring kinetics of ACE activity.