Kondo K, Sone H, Yoshida H, Toida T, Kanatani K, Hong Y M, Nishino N, Tanaka J
Central Laboratories of Key Technology, Kirin Brewery Co., Ltd., Gunma, Japan.
Mol Gen Genet. 1990 Mar;221(1):81-6. doi: 10.1007/BF00280371.
Arginine deiminase from Mycoplasma arginini was purified. The purified enzyme has a molecular weight of 46,000 daltons as determined by sodium dodecyl sulfate polyacrylamide gel electrophoresis. Its specific activity (20 units/mg protein) and amino acid composition showed a strong similarity to that of the Mycoplasma arthritidis arginine deiminase. The amino acid sequences of the N-terminal region and three internal peptides generated by enzymatic cleavage of the purified protein were determined. Using a synthetic oligonucleotide mixture complementary to part of the determined N-terminal amino acid sequence, the gene coding for arginine deiminase was isolated from a phage library. A nucleotide sequence of 2189 bp encoding the gene was determined. An open reading frame (ORF) contained the amino acid sequences corresponding to the determined N-terminal region and the three internal peptides of arginine deiminase. Thus it was concluded that this ORF encoded the arginine deiminase, a 385 amino acid polypeptide (mol.wt. 43,900 daltons). The three tryptophan residues in the sequenced peptides align with UGA codons in the nucleotide sequence, indicating that the nonsense codon UGA is used as a tryptophan codon in M. arginini.
对来自精氨酸支原体的精氨酸脱亚氨酶进行了纯化。通过十二烷基硫酸钠聚丙烯酰胺凝胶电泳测定，纯化后的酶分子量为46,000道尔顿。其比活性（20单位/毫克蛋白质）和氨基酸组成与关节炎支原体精氨酸脱亚氨酶具有很强的相似性。测定了纯化蛋白N端区域以及酶切产生的三个内部肽段的氨基酸序列。利用与所测定的N端氨基酸序列部分互补的合成寡核苷酸混合物，从噬菌体文库中分离出编码精氨酸脱亚氨酶的基因。测定了编码该基因的2189 bp核苷酸序列。一个开放阅读框（ORF）包含了与所测定的精氨酸脱亚氨酶N端区域以及三个内部肽段相对应的氨基酸序列。因此得出结论，该ORF编码精氨酸脱亚氨酶，这是一种由385个氨基酸组成的多肽（分子量43,900道尔顿）。测序肽段中的三个色氨酸残基与核苷酸序列中的UGA密码子对齐，表明在精氨酸支原体中无义密码子UGA被用作色氨酸密码子。
