Topic
Substrate (chemistry)
About: Substrate (chemistry) is a research topic. Over the lifetime, 35902 publications have been published within this topic receiving 740722 citations. The topic is also known as: enzyme substrate.
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TL;DR: A novel function for catalase is described potentially important in metabolism of endogenous substrates and in the action of carcinogens and chemopreventative agents.
137 citations
TL;DR: Crystalline plasma monoamine oxidase was prepared from beef blood by a procedure involving ammonium sulfate precipi- tation, diethylaminoethyl cellulose and hydroxylapatite chroma- tography, and was pink in color and showed an absorption maximum at about 480 mp.
137 citations
TL;DR: This synthetic substrate can be used in a rapid (2 min) and sensitive (10 ng of 31‐fold purified enzyme) spectrophotometric coupled enzyme assay for 2′,3′‐cyclic nucleotide 3′‐phosphohydrolase and the NADPH formed is measured by the increase in absorbance at 340 nm.
Abstract: – 2′,3′-Cyclic NADP has been prepared by cyclization of NADP at pH 6 in the presence of l-ethyl-(3-dimethylaminopropyl)-carbodiimide. The NADP derivative is readily hydrolyzed to NADP by the enzyme in brain and nerve that hydrolyzes 2′,3′-cyclic nucleotides to 2′-phospho esters. The Km for this substrate is the same as that for 2′,3′-cyclic AMP (0.22 mm) at pH 6 and 25°C. The two substrates are hydrolyzed by the phosphohydrolase at similar maximum velocities. The nicotinamide moiety in cyclic NADP thus has little effect on the enzyme-substrate interaction. This synthetic substrate can be used in a rapid (2 min) and sensitive (10 ng of 31-fold purified enzyme) spectrophotometric coupled enzyme assay for 2′,3′-cyclic nucleotide 3′-phosphohydrolase; in this assay the hydrolysis proceeds in the presence of glucose-6-phosphate dehydrogenase and its substrate and the NADPH formed is measured by the increase in absorbance at 340 nm. The assay is applicable to tissue extracts as well as to purified preparations of the enzyme. There is no interference from nucleases of the pancreatic RNase A type.
137 citations
136 citations
TL;DR: It is found that replacements of A sp248 affected glutamine turnover much more strongly than asparagine hydrolysis in variant N248A, and modeling studies suggested that the selective reduction of glutaminase activity is the result of small conformational changes that affect active‐site residues and catalytically relevant water molecules.
Abstract: The use of Escherichia coli asparaginase II as a drug for the treatment of acute lymphoblastic leukemia is complicated by the significant glutaminase side activity of the enzyme. To develop enzyme forms with reduced glutaminase activity, a number of variants with amino acid replacements in the vicinity of the substrate binding site were constructed and assayed for their kinetic and stability properties. We found that replacements of Asp248 affected glutamine turnover much more strongly than asparagine hydrolysis. In the wild-type enzyme, N248 modulates substrate binding to a neighboring subunit by hydrogen bonding to side chains that directly interact with the substrate. In variant N248A, the loss of transition state stabilization caused by the mutation was 15 kJ mol(-1) for L-glutamine compared to 4 kJ mol(-1) for L-aspartic beta-hydroxamate and 7 kJ mol(-1) for L-asparagine. Smaller differences were seen with other N248 variants. Modeling studies suggested that the selective reduction of glutaminase activity is the result of small conformational changes that affect active-site residues and catalytically relevant water molecules.
136 citations