Kinetics of heat inactivation of phenyl valerate hydrolases from hen and rat brain

Elsa Reiner, Cinda S. Davis, Bradley W. Schwab, Lawrence M. Schopfer, Rudy J. Richardson

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12 Scopus citations

Abstract

Heat inactivation was studied at 45, 50, 55, and 60° for all of the phenyl valerate hydrolases (PVase), including neurotoxic esterase (NTE) and inhibitor-resistant esterase (IRE), in homogenates of hen or rat brain or in preparations of hen brain microsomal membranes. Hen and rat brain homogenates were prepared in buffer (50 mM Tris/0.20 mM EDTA, pH 8.00, at 25°). Hen brain microsomes were suspended either in buffer or in aqueous dimethyl sulfoxide (DMSO, 40%, w v), or solubilized either in aqueous Triton X-100 (0.10%, w v) or in 40% ( w v) DMSO. Enzyme activities were measured at 37° using phenyl valerate as substrate. Each enzyme activity in all of the preparations exhibited biphasic heat inactivation kinetics. Apparent rate constants were calculated for the fast (kf) and slow (ks) reactions, along with the relative amounts of activity in each component (Af, As) expressed as percentages of the total activity. For a given preparation and temperature, respective values of kf or ks were similar for PVase, NTE, and IRE, with a mean kf ks, ratio of 52 across all preparations. Af and As, were a func of temperature. Mean values of the apparent activation energies (Ea) for all activities and preparations were 44 and 25 kcal/mol for the fast and slow inactivation reactions respectively. These results indicate that all phenyl valerate hydrolases in hen and rat brain undergo a common heat-induced structural change leading to loss of enzymic activity.

Original languageEnglish (US)
Pages (from-to)3181-3185
Number of pages5
JournalBiochemical Pharmacology
Volume36
Issue number19
DOIs
StatePublished - Oct 1 1987

ASJC Scopus subject areas

  • Biochemistry
  • Pharmacology

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