K30, H150, and H168 are essential residues for coordinating pyridoxal 5'-phosphate of O-acetylserine sulfhydrylase from Acidithiobacillus ferrooxidans.
O-acetylserine sulfhydrylase (OASS) is a key enzyme involved in the pathway of the cysteine biosynthesis. The gene of OASS from Acidithiobacillus ferrooxidans ATCC 23270 was cloned and expressed in E. coli, the soluble protein was purified by one-step affinity chromatography to apparent homogeneity. Colors and UV-vis scanning results of the recombinant protein confirmed that it was a pyridoxal 5'-phosphate (PLP)-containing protein. Sequence alignment and site-directed mutation of the enzyme revealed that the cofactor PLP is covalently bound in Schiff base linkage with K30, as well as the two residues H150 and H168 were the crucial residues for PLP binding and stabilization.
Zheng
Chunli
C
Donghua University, Shanghai, People's Republic of China. Zhengchunli1979@163.com
Nie
Li
L
Qian
Lin
L
Wang
Zhilou
Z
Liu
Guizhen
G
Liu
Jianshe
J
D042763:Acidithiobacillus / Q000201:enzymology* / Q000235:genetics / Q000378:metabolism; D000595:Amino Acid Sequence; D020134:Catalytic Domain; D003001:Cloning, Molecular; D003067:Coenzymes / Q000378:metabolism; D017124:Conserved Sequence; D003547:Cysteine Synthase / Q000737:chemistry* / Q000235:genetics* / Q000302:isolation & purification / Q000378:metabolism; D004926:Escherichia coli / Q000235:genetics; D008969:Molecular Sequence Data; D011485:Protein Binding; D011732:Pyridoxal Phosphate / Q000737:chemistry / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D012545:Schiff Bases; D012694:Serine / Q000031:analogs & derivatives / Q000378:metabolism
D003067:Coenzymes; D011994:Recombinant Proteins; D012545:Schiff Bases; D012694:Serine; D011732:Pyridoxal Phosphate; D003547:Cysteine Synthase; C043943:O-acetylserine