Bioproduction of L-Aspartic Acid and Cinnamic Acid by L-Aspartate Ammonia Lyase from Pseudomonas aeruginosa PAO1.
Appl Biochem Biotechnol
; 182(2): 792-803, 2017 Jun.
Article
em En
| MEDLINE
| ID: mdl-27988856
Aspartase (L-aspartate ammonia lyase, EC 4.3.1.1) catalyses the reversible amination and deamination of L-aspartic acid to fumaric acid which can be used to produce important biochemical. In this study, we have explored the characteristics of aspartase from Pseudomonas aeruginosa PAO1 (PA-AspA). To overproduce PA-AspA, the 1425-bp gene was introduced in Escherichia coli BL21 and purified. A 51.0-kDa protein was observed as a homogenous purified protein on SDS-PAGE. The enzyme was optimally active at pH 8.0 and 35 °C. PA-AspA has retained 56% activity after 7 days of incubation at 35 °C, which displays the hyperthermostablility characteristics of the enzyme. PA-AspA is activated in the presence of metal ions and Mg2+ is found to be most effective. Among the substrates tested for specificity of PA-AspA, L-phenylalanine (38.35 ± 2.68) showed the highest specific activity followed by L-aspartic acid (31.21 ± 3.31) and fumarate (5.42 ± 2.94). K m values for L-phenylalanine, L-aspartic acid and fumarate were 1.71 mM, 0.346 µM and 2 M, respectively. The catalytic efficiency (k cat/K m) for L-aspartic acid (14.18 s-1 mM-1) was higher than that for L-phenylalanine (4.65 s-1 mM-1). For bioconversion, from an initial concentration of 1000 mM of fumarate and 30 mM of L-phenylalanine, PA-AspA was found to convert 395.31 µM L-aspartic acid and 3.47 mM cinnamic acid, respectively.
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MEDLINE
Assunto principal:
Pseudomonas aeruginosa
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Aspartato Amônia-Liase
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Proteínas de Bactérias
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Cinamatos
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Ácido Aspártico
Idioma:
En
Ano de publicação:
2017
Tipo de documento:
Article