Cloning, expression and characterization of an aspartate aminotransferase gene from Lactobacillus brevis CGMCC 1306

Abstract

An aspartate aminotransferase (AATase) gene from Lactobacillus brevis CGMCC 1306 was cloned, which contains a 1182-bp open reading frame coding for 393 amino acids (41.43 kDa). When expressed in Escherichia coli BL21 (DE3), the recombinant AATase was purified and subsequently characterized. The recombinant AATase can catalyse the conversion of L-Asp to L-Glu, and the k cat / K m was determined to be 25.5 (mmol/L) ¡1 s ¡1 for L-Asp and 207.8 m(mol/L) ¡1 s ¡1 for a-ketoglutarate. With optimum temperature as 25 8C, the AATase may be a novel and special psychrophilic enzyme which exhibited a good thermal stability below 55 8C. The conserved active site residue of AATase was identified as Lys237 by phylogenetic analysis. Secondary structure of the enzyme includes a-helix (39.2%), b-sheet (5.5%), b-turn (8.8%), and random coil (36.5%) by circular dichroism spectral analysis. Phase diagram for the fluorescence data analysis showed that guanidinium chloride-induced unfolding of AATase involved at least one intermediate.

Volatile Organic Compounds (VOCs) Produced by Levilactobacillus brevis WLP672 Fermentation in Defined Media Supplemented with Different Amino Acids

“…Brevis ATCC 367 [ 53 ] and Lev. Brevis CGMCC 1306 [ 62 ]. Hence, it seems more likely that, in the current study, AspAT in LB672 carried out the transamination reaction of Phe rather than the ArAT enzyme.…”

Engineering the bacterial endophyte Burkholderia pyrrocinia JK-SH007 for the control of lepidoptera larvae by introducing the cry218 genes of Bacillus thuringiensis

“…Two pairs of specific primers (Supplemental Table 1S) were designed respectively according to the DNA sequences of the Cry218-pht304 plasmid and PHKT2 recipient expression vector [10]. These amplified DNA fragments were mixed at a ratio of 1:3 (100 ng vector to 300 ng DNA fragments), and transformed into E. coli XL-10 Gold cells [11].…”