Functional identification of the gene locus (ncg12319 and characterization of catechol 1,2-dioxygenase in Corynebacterium glutamicum.
Corynebacterium glutamicum assimilated phenol, benzoate, 4-hydroxybenzoate p-cresol and 3,4-dihydroxybenzoate. Ring cleavage was by catechol 1,2-dioxygenase when phenol or benzoate was used and by protocatechuate 3,4-dioxygenase when the others were used as substrate. The locus ncg12319 of its genome was cloned and expressed in Escherichia coli. Enzyme assays showed that ncg12319 encodes a catechol 1,2-dioxygenase. This catechol 1,2-dioxygenase was purified and accepted catechol, 3-, or 4-methylcatechols, but not chlorinated catechols, as substrates. The optimal temperature and pH for catechol cleavage catalyzed by the enzyme were 30 degrees C and 9, respectively, and the Km and Vmax were determined to be 4.24 micromol l(-1) and 3.7 micromol l(-1) min(-1) mg(-1) protein, respectively.
Shen
Xi-Hui
XH
Institute of Microbiology, Chinese Academy of Sciences, Beijing 100080, PR China.
Liu
Zhi-Pei
ZP
Liu
Shuang-Jiang
SJ
D000595:Amino Acid Sequence; D050548:Catechol 1,2-Dioxygenase; D002874:Chromosome Mapping; D003001:Cloning, Molecular / Q000379:methods; D003352:Corynebacterium / Q000201:enzymology* / Q000235:genetics*; D049308:Dioxygenases / Q000737:chemistry* / Q000235:genetics* / Q000378:metabolism; D004789:Enzyme Activation; D004795:Enzyme Stability; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics; D020869:Gene Expression Profiling; D008969:Molecular Sequence Data; D008970:Molecular Weight; D011487:Protein Conformation; D011994:Recombinant Proteins / Q000737:chemistry / Q000378:metabolism; D017386:Sequence Homology, Amino Acid
D011994:Recombinant Proteins; D049308:Dioxygenases; D050548:Catechol 1,2-Dioxygenase