KINETICS OF THE REACTION OF RENIN WITH NINE SYNTHETIC PEPTIDE SUBSTRATES

A number of peptides have been synthesized which represent portions of the tetradecapeptide renin substrate molecule, and which contain the hydrolyzable leu-leu bond. An automatic chemical method for determination of the velocity of the reaction of renin with these compounds was developed. Application of the method at several levels of substrate concentration permitted construction of Lineweaver-Burk plots, and calculation of Michaelis constants (K_m) and maximal velocities (V_max). The results show that the maximum affinity of the enzyme (lowest K_m) for substrate is achieved only with the full tetradecapeptide molecule (asp¹-arg²-val³-tyr⁴-ileu⁵-his⁶-pro⁷-phe⁸-his⁹-leu¹⁰-leu¹¹-val¹²-tyr¹³-ser¹⁴). Removal of asp¹ and arg² from the N-terminal increases the K_m eight-fold. Further, moderate increase in K_m occurs when the next amino acids, val³, tyr⁴ and ileu⁵, are removed. The further removal of his⁶ results in a marked reduction in the V_max. Removal of ser¹⁴ from the C-terminal of the nonapeptide his⁶-pro⁷-phe⁸-his⁹-leu¹⁰-leu¹¹-val¹²-tyr¹³-ser¹⁴ does not greatly affect the K_m nor the V_max. Further removal of tyr¹³ from this compound results in complete loss of substrate activity. It is suggested that the compounds his⁶-pro⁷-phe⁸-his⁹-leu¹⁰-leu¹¹-val¹²-tyr¹³-ser¹⁴ or his⁶-pro⁷-phe⁸-his⁹-leu¹⁰-leu¹¹-val¹²-tyr¹³ might be used as substrates for the chemical assay and standardization of renin.