Redesign of Choline Acetyltransferase Specificity by Protein Engineering
نویسندگان
چکیده
منابع مشابه
Redesign of carnitine acetyltransferase specificity by protein engineering.
In eukaryotes, L-carnitine is involved in energy metabolism by facilitating beta-oxidation of fatty acids. Carnitine acetyltransferases (CrAT) catalyze the reversible conversion of acetyl-CoA and carnitine to acetylcarnitine and free CoA. To redesign the specificity of rat CrAT toward its substrates, we mutated Met564. The M564G mutated CrAT showed higher activity toward longer chain acyl-CoAs:...
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Immunohistochemical localization of choline acetyltransferase (ChAT) in cholinergic neurons has been difficult to achieve because of problems encountered in producing specific antisera. Here we describe the production and characterization of several distinct monoclonal antibodies to ChAT. Each of the monoclonal antibodies exhibits one of three general patterns of cross-species reactions; one pa...
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Four monoclonal antibodies were obtained to rat brain choline acetyltransferase (CAT). The enzyme was purified 95,000-fold from rat brain by precipitation with acetic acid at pH 4.5, fractionation with 40 to 60% (NH4)2SO4, CM-Sephadex chromatography, and affinity column chromatography on agarose-hexane-coenzyme A. The enzyme preparation was applied to the affinity column in the presence of 10 m...
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Choline acetyltransferase (EC 2.3.1.6) catalyzes the reversible transfer of the acetyl group from acetyl coenzyme A to choline. Previous studies are consistent with the idea that an active site sulfhydryl group reacts with acetyl coenzyme A to form an acetyl-thioenzyme intermediate and coenzyme A (ROSKOSKI, R., JR. (1973) Biochemistry 12, 3709). Choline then reacts with the acetyl-enzyme to for...
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The localization, purification, and some enzymatic properties of choline acetyltransferase from rat brain were studied. Most assays were performed with a specific radiometric micromethod. Solubilization of the enzyme was examined after homogenization of cerebral cortices by means which disintegrate nerve endings. In isotonic KC1 the enzyme was recovered in solution. In more dilute KC1 the enzym...
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ژورنال
عنوان ژورنال: Journal of Biological Chemistry
سال: 1998
ISSN: 0021-9258
DOI: 10.1074/jbc.273.38.24465