A novel serine hydroxymethyltransferase from Arthrobacter nicotianae: characterization and improving catalytic efficiency by rational design.
BACKGROUND
Serine hydroxymethyltransferase (SHMT) is the key enzyme in L-serine enzymatic production, suggesting the importance of obtaining a SHMT with high activity.


RESULTS
Here, a novel SHMT gene, glyA, was obtained through degenerate oligonucleotide-primed PCR and encoded a novel SHMT with 54.3% similarity to the known SHMT from Escherichia coli. The obtained protein AnSHMT showed the optimal activity at 40 °C and pH 7.5, and was more stable in weakly alkali conditions (pH 6.5-8.5) than Hyphomicrobium methylovorum's SHMT (pH 6.0-7.5), In order to improve the catalytic efficiency of the wild type, the site-directed mutagenesis based on sequences alignment and bioinformatics prediction, was used and the catalytic efficiency of the mutant I249L was found to be 2.78-fold higher than that of the wild-type, with the replacement of isoleucine by leucine at the 249 position.


CONCLUSIONS
This research provides useful information about the interesting site, and the application of DOP-PCR in cloning a novel glyA gene.
State Key Laboratory of Agricultural Microbiology, College of Life Science and Technology, Huazhong Agricultural University, Wuhan, 430070, P. R. China. tianya416@126.com.
Wei
W
Jiang
State Key Laboratory of Agricultural Microbiology, College of Life Science and Technology, Huazhong Agricultural University, Wuhan, 430070, P. R. China. chenlin19890717@163.com.
Lin
L
Chen
College of Biotechnology and Pharmaceutical Engineering, Nanjing Tech University, Nanjing, 211800, P. R. China. hunan@njtech.edu.cn.
Nan
N
Hu
State Key Laboratory of Agricultural Microbiology, College of Life Science and Technology, Huazhong Agricultural University, Wuhan, 430070, P. R. China. 984023989@qq.com.
Shaohui
S
Yuan
State Key Laboratory of Agricultural Microbiology, College of Life Science and Technology, Huazhong Agricultural University, Wuhan, 430070, P. R. China. 985927577@qq.com.
Bin
B
Li
State Key Laboratory of Agricultural Microbiology, College of Life Science and Technology, Huazhong Agricultural University, Wuhan, 430070, P. R. China. lzd@mail.hzau.edu.cn.
Ziduo
Z
Liu
D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D001173:Arthrobacter / Q000737:chemistry / Q000145:classification / Q000201:enzymology* / Q000235:genetics; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D055162:Biocatalysis; D003001:Cloning, Molecular; D004795:Enzyme Stability; D012696:Glycine Hydroxymethyltransferase / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D007700:Kinetics; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D010802:Phylogeny; D015202:Protein Engineering; D016415:Sequence Alignment
D001426:Bacterial Proteins; D012696:Glycine Hydroxymethyltransferase