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B6db references: 4917239

type Journal Article
authors Dowhan W Jr, Snell EE.
title D-serine dehydratase from Escherichia coli. II. Analytical studies and subunit structure
journal J Biol Chem
sel selected
ui 4917239
year (1970)
volume 245
number 18
pages 4618-28
abstract An improved procedure which permits large scale preparation of crystalline D-serine dehydratase (EC from Escherichia coli is described. The homogeneous holoenzyme has an extinction coefficient (E 1%) of 10.5 in potassium phosphate, pH 6.5 to 7.8, a specific activity of 300, and contains a single peptide chain with a molecular weight near 45,500 as indicated by sedimentation equilibrium measurements of either native or reduced and carboxyxnethylated enzyme, acrylamide gel electrophoresis in sodium dodecyl sulfate, quantitative determination of the N-terminal amino acid (methionine), and the results of tryptic peptide mapping. One combining site for pyridoxal-P is present per molecule as determined by direct analysis of the holoenzyme for pyridoxal-P, by spectrophotometric titration of the apoenzyme with pyridoxal-P, and by isolation of e-N-pyridoxyllysine from hydrolysates of the sodium borohydride-reduced holoenzyme. The enzyme contains 5 half-cystine residues and no disulfide linkages. Only one of the five sulfhydryl groups is titrated by 5,5-dithio-bis(Z-nitrobenzoic acid) (DTNB) in the holoenzyme; in the apoenzyme three -SH groups are titrated by either p-chloromercuribenzoate or DTNB. These three groups react with DTNB at markedly different rates (0.54, 0.081, and 0.0033 min-l) under the conditions tested. Individually, none of the three -SH groups appears necessary for activity; however, reaction of two or more with DTNB inactivates the apoenzyme completely by preventing its reassociation with pyridoxal-P.
last changed 2009/06/30 14:57

B6db references