Purification, cDNA cloning and expression of human NG,NG-dimethylarginine dimethylaminohydrolase.
cDNA encoding N(G),N(G)-dimethylarginine dimethylaminohydrolase from rat kidney had been cloned [Kimoto, M., Sasakawa, T., Tsuji, H., Miyatake, S., Oka, T., Nio, N. & Ogawa, T. (1997) Biochim. Biophys. Acta 1337, 6-10]. The enzyme hydrolyzes N(G),N(G)-dimethyl-L-arginine and N(G)-monomethyl-L-arginine, which are known as endogenous inhibitors for the nitric oxide-generating system. In the present study, human N(G),N(G)-dimethylarginine dimethylaminohydrolase has been purified to homogeneity from liver and characterized. The cDNA clone encoding human N(G),N(G)-dimethylarginine dimethylaminohydrolase was isolated from a human kidney lambda gt10 library using a probe prepared from a plasmid containing the entire coding region of rat N(G),N(G)-dimethylarginine dimethylaminohydrolase. Its open reading frame encoded a protein of 285 amino acids with a molecular mass of 31,121 Da. The deduced amino acid sequence exhibits 93% identity with that of rat. The cDNA was expressed as a fusion protein in Escherichia coli and the recombinant protein exhibited enzyme activity which is the same as that of natural enzyme.
Kimoto
M
M
Department of Nutritional Science, Faculty of Health and Welfare Science, Okayama Prefectural University, Soja, Japan. kimoto@fhw.oka-pu.ac.jp
Miyatake
S
S
Sasagawa
T
T
Yamashita
H
H
Okita
M
M
Oka
T
T
Ogawa
T
T
Tsuji
H
H
D000581:Amidohydrolases; D000595:Amino Acid Sequence; D001483:Base Sequence; D003001:Cloning, Molecular; D006801:Humans; D006867:Hydrolases / Q000235:genetics*; D007700:Kinetics; D008099:Liver / Q000201:enzymology*; D008969:Molecular Sequence Data; D012333:RNA, Messenger / Q000235:genetics; D011994:Recombinant Proteins / Q000235:genetics; D017422:Sequence Analysis, DNA; D017386:Sequence Homology, Amino Acid; D014357:Trypsin / Q000378:metabolism
D012333:RNA, Messenger; D011994:Recombinant Proteins; D006867:Hydrolases; D014357:Trypsin; D000581:Amidohydrolases; C059999:dimethylargininase