tailieunhanh - Báo cáo khoa học: Cloning, expression and characterization of a new aspartate aminotransferase from Bacillus subtilis B3

In the present study, we report the identification of a new gene from the Bacillus subtilisB3 strain (aatB3), which comprises 1308 bp encoding a 436 amino acid protein with a monomer molecular weight of kDa. Phylo-genetic analyses suggested that this enzyme is a member of the Ib subgroup of aspartate aminotransferases | 1FEBS Journal Cloning expression and characterization of a new aspartate aminotransferase from Bacillus subtilis B3 Hui-Jun Wu1 Yang Yang1 z Shuai Wang2 z Jun-Qing Qiao1 Yan-Fei Xia1 Yu Wang1 Wei-Duo Wang1 Sheng-Feng Gao1 Jun Liu1 Peng-Qi Xue1 and Xue-Wen Gao1 1 Department of Plant Pathology College of Plant Protection Nanjing AgriculturalUniversity Key Laboratory of Monitoring and Management of Crop Diseases and Pest Insects Ministry of Agriculture China 2 Key Laboratory of Synthetic Biology Institute of Plant Physiology and Ecology Shanghai Institutes for BiologicalSciences Chinese Academy of Sciences China Keywords aspartate aminotransferase Bacillus subtilis conserved active residues kinetic parameters protein sequence analysis In the present study we report the identification of a new gene from the Bacillus subtilis B3 strain aatB3 which comprises 1308 bp encoding a 436 amino acid protein with a monomer molecular weight of kDa. Phylogenetic analyses suggested that this enzyme is a member of the Ib subgroup Correspondence . Gao Department of Plant Pathology College of Plant Protection Nanjing AgriculturalUniversity Key Laboratory of Monitoring and Management of Crop Diseases and Pest Insects Ministry of Agriculture Nanjing 210095 China Fax 86 25 84395268 Tel 86 25 84395268 E-mail gaoxw@ of aspartate aminotransferases AATs EC although it also has conserved active residues and thermostability characteristic of Ia-type AATs. The Asp232 Lys270 and Arg403 residues of AATB3 play a key role in transamination. The enzyme showed maximal activity at pH and 45 C had relatively high activity over an alkaline pH range pH and was stable up to 50 C. AATB3 catalyzed the transamination of five amino acids with L-aspartate being the optimal substrate. The Km values were determined to be mM for L-aspartate mM for a-ketoglutarate mM for L-glutamate and mM for oxaloacetate. A 32-residue N-termi-nal amino acid sequence of this