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B6db references: 94115184

type Journal Article
authors Yamauchi, A.; Stijntjes, G. J.; Commandeur, J. N.; Vermeulen, N. P.
title Purification of glutamine transaminase K/cysteine conjugate beta-lyase from rat renal cytosol based on hydrophobic interaction HPLC and gel permeation FPLC
journal Protein Expr Purif
ui 94115184
year (1993)
volume 4
number 6
pages 552-62.
keywords Amino Acids/analysis
abstract Cysteine conjugate beta-lyase (beta-lyase, EC was purified to homogeneity from rat renal cytosol using a new and highly efficient method, based on C3-hydrophobic interaction (HI) high-performance liquid chromatography (HPLC) in combination with gel permeation fast protein liquid chromatography. The purity of the enzyme was judged from SDS-PAGE and C18-reversed-phase HPLC. The beta-lyase was estimated to be a homodimer consisting of a 47,400-Da subunit with absorption maxima at 280 and 420-430 nm. The specific activity of the purified beta-lyase toward S-(1,2-dichlorovinyl)-L-cysteine (1,2-DCVC) in the presence of alpha-keto-gamma-methiolbutyric acid (KMB) was 6.4 mumol/min/mg protein, which is by far the highest value so far reported. Kinetic analysis of 1,2-DCVC metabolism by the enzyme in the presence of KMB gave Km and Vmax values of 0.33 mM and 8.4 mumol/min/mg protein, respectively. No significant activity of the purified enzyme was detectable with S-2-benzothiazolyl-L-cysteine up to 2 mM. The purified enzyme also had glutamine transaminase K activity (EC as assayed with phenylalanine and KMB as substrates. This specific activity was 16.0 mumol/min/mg. Amino acid analysis of the purified beta-lyase was carried out and was found to be closely similar to the amino acid composition of five other pyridoxal phosphate (PLP)- containing amino acid amino-transferases. This suggests that glutamine transaminase K/cysteine conjugate beta-lyase is a typical member of the PLP-containing aminotransferase group.
last changed 2002/11/12 16:17

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