Vitamin K1 dependent carboxylase: beta-carboxylation of t-butyloxycarbonylaspartic acid alpha-benzyl ester.
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Animals↗
Biomedical subjects
S E Hamilton
Publications and source records attributed to S E Hamilton.
The nature and reactivity of the "essential" thiol in rabbit muscle creatine kinase III (EC 2.7.3.2).
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Animals↗
Purification and properties of a carboxylesterase from the liver of tiger shark (Galeocerdo cuvier).
A procedure is described for the purification of a carboxylesterase from shark liver, using a chloroform-acetone powder prepared from the liver as the starting material. The yield of purified enzyme is approximately 50 mg from 530 g of chloroform-acetone powder. The preparation is electrophoretically homogeneous. Active-site titrations with paraoxon gave an equivalent weight of approximately 83 000. The molecular weight, found from sedimentation equilibrium experiments, is approximately 80 000. There is no evidence of any association or dissociation of this species. The enzyme shows a marked preference for aryl esters over alkyl esters, in contrast to other carboxylesterases so far studied. The amino acid composition of the purified enzyme is reported.
Amino Acids↗
Organophosphate inhibitors: the reactions of bis(p-nitrophenyl) methyl phosphate with liver carboxylesterases and alpha-chymotrypsin.
Bis(p-nitrophenyl) methyl phosphate (BNMP) has been tested as a spectrophotometric titrant for a group of serine hydrolases. Bis(p-nitrophenyl) methyl phosphate reacts rapidly with liver carboxylesterases from chicken, sheep, and horse, and more slowly with alpha-chymotrypsin, releasing 2 mol of p-nitrophenol per active site titrated, and producing a phosphorylated enzyme very stable to dephosphorylation. However, pig liver carboxylesterase produces 2.2 mol of p-nitrophenol per active site titratedmreaction of pig and chicken liver carboxylesterases with bis(p-nitrophenyl) [3H]methyl [32P]phosphate clarified this differencemone molecule of the chicken enzyme reacts with one molecule of bis(p-nitrophenyl) methyl phosphate, releasing both p-nitrophenol residues, and resulting in an inhibited enzyme with one phosphorus atom and one methyl group covalently bound. Pig enzyme reacts rapidly, forming (presumably) methyl p-nitrophenyl phosphoryl-carboxylesterasemthis further reacts, concurrently producing methyl phosphoryl-carboxylesterase plus p-nitrophenol, or free enzyme plus methyl p-nitrophenyl phosphate, in the ratio of about 5 : 1 at pH 7.55. The free enzyme produced undergoes further reaction with bis(p-nitrophenyl) methyl phosphate until all the carboxylesterase is inhibited.
Animals↗
Carboxylesterases (EC 3.1.1). Purification and titration of chicken, sheep, and horse liver carboxylesterases.
Chicken, sheep, and horse liver carboxylesterases have been purified by procedures involving ammonium sulfate fractionation, ion-exchange chromatography and gel filtration on Sephadex. The actual yields of the procedures described were as follows: chicken, 1 g from 2 kg of liver powder (chloroform-acetone); sheep, 200 mg from 400 g of powder (chloroform-acetone); horse, 230 mg from 800 g of powder (acetone). The purified enzymes are free of non-carboxyl-esterase protein as shown by gel electrophoresis, although they do contain electrophoretic variants. The equivalent weight of the chicken enzyme is 67,000 based on titration with p-nitrophenyl diethyl phosphate or bis(p-nitrophenyl) phosphate, whereas those of the sheep and horse enzymes are similar to 69,500 and similar to 70,000, respectively, based on titration with p-nitrophenyl dimethylcarbamate.
Ammonium Sulfate↗
Carboxylesterases (EC 3.1.1). A comparison of some kinetic properties of horse, sheep, chicken, pig, and ox liver carboxylesterases.
A comparative study of the kinetic be,avior of horse, sheep, chicken, pig, and ox liver carboxylesterases is reported. The enzymes exhibit similar specificites towards a series of phenyl esters in which the acyl group is varied, and towards a series of butyrate esters in which the alcohol group is varied. Non-Michaelis-Menten kinetics are exhibited by the horse enzyme in the hydrolysis of methyl and ethyl butyrates, and by the pig enzyme with ethyl butyrate. Each enzyme exhibits inhibition by one or more substrates. A simple scheme which accounts for both activation and inhibition is discussed. pH-k(cat) profiles for the horse and chicken liver carboxylesterase-catalyzed hydrolyses of phenyl butyrate demonstrate dependencies on pK(a)S of 4.75 and 5.0, respectively.
Animals↗
Molecular mechanisms for the regulation of the expression and function of muscarinic acetylcholine receptors.
The regulation of muscarinic acetylcholine receptor expression and function was investigated in cultured cells and in knockout mice. Muscarinic agonist exposure causes m2 receptor desensitization and sequestration and decreases the expression of cardiac potassium channels. The expression of m2 receptors in chick retina is regulated by a developmentally regulated secreted factor. Mice lacking the m1 receptor exhibit a loss of muscarinic regulation of M-current potassium channel activity and pilocarpine-induced seizures.
Animals↗