|
type |
Journal Article |
authors |
Bhattacharjee, M. K.; Snell, E. E. |
title |
Pyridoxal 5'-phosphate-dependent histidine decarboxylase. Mechanism of inactivation by alpha-fluoromethylhistidine |
journal |
J Biol Chem |
Activity |
4.1.1.22 |
ui |
90216689 |
year |
(1990) |
volume |
265 |
number |
12 |
pages |
6664-8. |
| |
keywords |
Carboxy-Lyases/*antagonists & inhibitors |
abstract |
Mechanism-based inactivation of pyridoxal phosphate-dependent histidine decarboxylase by (S)-alpha-(fluoromethyl)histidine was studied. The molar ratio of inactivator to enzyme subunit required for complete inactivation increased from 1.63 at 10 degrees C to 3.00 at 37 degrees C. Two inactivation products were isolated by chromatographic fractionation of the reaction mixture and identified by NMR spectroscopy as 1-(4-imidazolyl)-3(5'-P-pyridoxylidene) acetone (I), the adduct formed between pyridoxal phosphate and inactivator, and 1-(4- imidazolyl) acetone (II), an intermediate compound formed during inactivation. Formation of these two products supports a previously proposed mechanism of inactivation (Hayashi, H., Tanase, S., and Snell, E. E. (1986) J. Biol. Chem. 261, 11003-11009), with minor modifications. A precursor of I was linked covalently to the enzyme by NaBH4 reduction if the reaction was carried out immediately after inactivation, before development of the 403 nm peak of I. A mutant histidine decarboxylase (S322A) in which Ser-322 was changed to Ala was also inactivated by alpha-fluoromethylhistidine demonstrating that Ser- 322 is not essential for inactivation even though it is close to the active site and is derivatized by borohydride reduction of the inactivated wild-type enzyme. Following inactivation, both the wild- type and the S322A mutant enzyme could be partially reactivated by prolonged dialysis against buffer. |
last changed |
2002/11/12 16:17 |
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