Download e-book for iPad: Affinity labeling by Nathan P. Kaplan, Nathan P. Colowick, William B. Jakoby,

By Nathan P. Kaplan, Nathan P. Colowick, William B. Jakoby, Meir Wilchek

ISBN-10: 0121819469

ISBN-13: 9780121819460

The significantly acclaimed laboratory normal, Methods in Enzymology, is without doubt one of the such a lot hugely revered courses within the box of biochemistry. when you consider that 1955, each one quantity has been eagerly awaited, usually consulted, and praised by means of researchers and reviewers alike. The sequence comprises a lot fabric nonetheless suitable this present day - really a vital e-book for researchers in all fields of lifestyles sciences

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6W. Birchmeier, P. E. Zaoralek, and P. Christen, Biochemistry 12, 2874 (1973). eK. J. Wilson, W. Birchmeier, and P. Christen, Eur. J. Biochem. 41, 471 (1974). < 0 Z ~ o ¢; ~a <'-. ©~,, ~ ~N + ° o o • °~ 44 GENERAL METHODOLOGY ~] enzyme the rate of modification of Cys-390 is at a minimum. The rate increases as the enzyme passes through the consecutive enzyme-substrate intermediates and decreases again when the free pyridoxamine form is reached. The rate of Cys-390 modification is fastest when, in the presence of a substrate pair (glutamate plus 2-ketoglutarate or aspartate plus oxaloacetate), covalent enzyme-substrate intermediates are formed.

Cassio, F. LeMoine, J. P. Waller, E. Sandrin, and R. A. Boissonas, Biochemistry 6, 827 (1967). ~'Y. C. Lee, Biochem. Biophys. Res. Commun. 35, 161 (1969). ~D. F. Wentworth and R. Wolfenden, Biochemistry 13, 4715 (1975). [2] TRANSITION S T A T E ANALOGS 27 Inhibition: hi E(H) + h-1 k2 ~ H + E(H) I-~O Normal reaction: E(H) R + - OH H~O m OH H OH Consistent with this hypothesis, rates of onset and rel'ease from inhibition were retarded in D20, but to quite different extents, K~ itself showing a pronounced solvent isotope effect.

The amino acid shows color changes of yellow to gray to purple with ninhydrin. This color change is characteristic of simple fl, ~-unsaturated amino acids. Vinyl glycine can be quantitatively reduced with hydrogen to afford DL-2-aminobutanoic acid. 4 mg of p-nitrophenyl chloroformate is added dropwise to 10 ml of a cold, stirred solution of diazomethane in ether (at least 4 mmoles of diazomethane, or a 16-fold excess, are used). The solution is allowed to stand overnight at 4 °. Excess diazomethane is removed in a stream of nitrogen, and the ether is evaporated under reduced pressure.

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Affinity labeling by Nathan P. Kaplan, Nathan P. Colowick, William B. Jakoby, Meir Wilchek

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