Putidaredoxin还原酶和Putidaredoxin。克隆、序列测定和异源表达的蛋白质。
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彼得森是的,Lorence MC, Amarneh B
Putidaredoxin还原酶和Putidaredoxin。克隆、序列测定和异源表达的蛋白质。
J生物化学杂志。1990年4月15日,265 (11):6066 - 73。
- PubMed ID
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2180940 (在PubMed]
- 文摘
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樟脑的氧化细胞色素p - 450凸轮需要黄素蛋白的参与,putidaredoxin还原酶,和一个iron-sulfur蛋白质,putidaredoxin,调解的转移电子从NADH为氧激活p - 450。一双2.2千碱基BamHI-StuI从整个细胞的DNA片段camphor-grown假单胞菌putida已被克隆和测序。翻译的序列显示两个开放阅读框架代码putidaredoxin还原酶和putidaredoxin。putidaredoxin而言,翻译序列匹配发表序列(田中,M。Haniu, M。Yasunobu, k . T。杜,K。Gunsalus, i c(1974)生物。249年化学,3689 - 3701)除了氨基酸之一。密码子的使用在这些蛋白质,像其他假单胞菌的蛋白质,强烈的偏见G + C在第三个核苷酸。发现了一个潜在的转录终止网站3 ' putidaredoxin编码区。“FAD-binding”氨基酸共识序列,在其他黄素蛋白,被发现在putidaredoxin还原酶开始残渣11和第二个出现的序列氨基酸156年开始被发现。 The second sequence could represent the NAD-binding site. The regions encoding putidaredoxin reductase and putidaredoxin were subcloned and independently expressed in Escherichia coli at the level of 0.4 and 4.8 mg of enzymatically active protein/g wet weight of cells, respectively. Site-directed mutagenesis was used to change the rare start codon, GTG, of putidaredoxin reductase to ATG which resulted in an 18-fold increase in the level of expression of this protein to 7.4 mg/g wet weight of cells. The construction of these two clones, which express these important proteins, will facilitate studies of their interaction with each other and with P-450cam.