Reference

Alternate substrates of dopamine b-hydroxylase

Alternate substrates of dopamine b-hydroxylase. I. Kinetic investigations of benzyl cyanides as substrates and inhibitors. Colombo, Giovanna; Rajashekhar, B.; Giedroc, David P.; Villafranca, Joseph J. (Dep. Chem., Pennsylvania State Univ., University Park, PA 16802, USA). J. Biol. Chem. 51-67-2 and 19924-43-7 are cas registry numbers. These chemicals are also mentioned in this article., 259(3), 1593-600 (English) 1984. CODEN: JBCHA3. ISSN: 0021-9258. DOCUMENT TYPE: Journal CA Section: 7 (Enzymes) A series of benzylcyanide analogs were studied as substrates and inhibitors of dopamine b-hydroxylase (I). p-Hydroxybenzylcyanide was previously shown to be a suicide substrate of I. The apparent max. velocity (Vmax) values for substituted benzylcyanide analogs decreased in the order: p-OH > m-OH > H >> p-OCH3, m-OCH3; the m-OH, m-OCH3, and p-OCH3 analogs were competitive inhibitors vs. tyramine in initial velocity studies. The Vmax values for tyramine and p-hydroxybenzylcyanide were nearly identical at satg. O2 and ascorbate (pH 5.0, 37%), but the Michaelis const. (Km) for O2 was 0.14 and 2.8 mM, resp., with tyramine and p-hydroxybenzylcyanide. Studies of the pH dependence of log Vmax/Km for tyramine showed 2 pKa values of 5.2 and 5.8, whereas for m-hydroxybenzylcyanide the values were 5.3 and 5.9. The log Vmax profile showed 1 pKa of 5.9 with tyramine as substrate. Thus, nearly identical enzymic groups are involved in binding and(or) catalysis with these 2 substrates. All of the benzylcyanide analogs were suicide inactivators of I. With m-hydroxybenzylcyanide, the partition between catalysis and inactivation (kcat/kinact) changed from ~600 to ~17 as the pH varied from 5.0 to 6.7. The log kinact vs. pH profile showed 1 pKa value of 6.0, suggesting that an enzymic group must be deprotonated for maximal inactivation. Cu was essential for the suicide inactivation of I by benzylcyanides and kinetic studies of partially inhibited I (~50%) showed that inactive enzyme mols. were completely inactive. .