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B6db references: 16847601

type Journal Article
authors Wang NC, Lee CY.
title Molecular cloning of the aspartate 4-decarboxylase gene from Pseudomonas sp. ATCC 19121 and characterization of the bifunctional recombinant enzyme
journal Appl Microbiol Biotechnol
Activity 4.1.1.12
Family 4.1.1.12
sel selected
ui 16847601
year (2006)
volume 73
number 2
pages 339-48
 
abstract L-Aspartate 4-decarboxylase (Asd) is a major enzyme used in the industrial production of L-alanine. Its gene was cloned from Pseudomonas sp. ATCC 19121 and characterized in the present study. The 1,593-bp asd encodes a protein with a molecular mass of 59,243 Da. The Asd from this Pseudomonas strain was considerably homologous to other Asds and aminotransferases, and has evolved independently of these enzymes from gram-positive microbes. Productivity rate of the C-terminal His-tagged fusion Asd was at 33 mg/l of Escherichia coli transformant culture. The kinetic parameters K (m) and V (max) of the fusion protein were 11.50 mM and 0.11 mM/min, respectively. Gel filtration analysis demonstrated that Asd is a dodecamer at pH 5.0 while 4.4 % of the recombinant protein dissociated into dimer when the pH was increased to 7.0. Asd exhibited its maximum activity at pH 5.0 and specific activity of 280 U/mg, and remained stable over a broad range of pH. The optimum temperature for Asd reaction was 45 degrees C, and 92 % of the activity remained when the enzyme was incubated at 40 degrees C for 40 min. This enzyme did not have any preferred divalent cation for catalysis. The recombinant Asd also exhibited aminotransferase activity when D,L-Asp, L-Glu, L-Gln, and L-Ala were utilized as substrates. However, the decarboxylation activity of L-aspartate was 2,477 times higher than its aminotransferase activity. The present study is the first investigation on the important biochemical properties of the purified recombinant Asd.
last changed 2007/12/21 17:48

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