Nakatsu C H, Straus N A, Wyndham R C
Institute of Biology, Carleton University, Ottawa, Ontario, Canada.
Microbiology (Reading). 1995 Feb;141 ( Pt 2):485-95. doi: 10.1099/13500872-141-2-485.
The nucleotide sequence of the 3-chlorobenzoate 3,4-dioxygenase genes, designated cbaAB, from the transposon Tn5271 was determined. The function of the two sequenced open reading frames was evaluated by mutagenesis and expression in vivo to show that the cbaA and cbaB genes code for dioxygenase and reductase proteins, respectively. Comparison of the deduced amino acid sequences of the cbaAB genes with sequences for other oxygenases revealed a clearly defined lineage among the class IA oxygenases that shows several unique features. This lineage includes phthalate 4,5-dioxygenase (pht23), and based on the available NH3-terminal sequence of component A, also includes 4-sulphobenzoate 3,4-dioxygenase. Vanillate demethylase, encoded by the vanAB genes and formally a monooxygenase enzyme catalysing an oxidative demethylation, is also included in this lineage. The terminal chlorobenzoate dioxygenase (CbaA) component is characterized by a conserved Rieske-type [2Fe-2S]R ligand centre. The reductase component (CbaB) contains a plant-type ferredoxin [2Fe-2S]Fd, FMN-isoalloxazine and NAD-ribose-binding domains and the orientation of these domains is conserved in all known class IA reductases. These results support the hypothesis that alternative fusions of the electron transfer modules of the reductases arose early in the divergence of oxygenase systems. The over-riding evolutionary constraint acting on the divergence of the class IA oxygenases would appear to be the requirement for a carboxyl group para to the site of oxygen insertion into the aromatic ring.
测定了转座子Tn5271中3-氯苯甲酸3,4-双加氧酶基因（命名为cbaAB）的核苷酸序列。通过诱变和体内表达评估了两个测序的开放阅读框的功能，结果表明cbaA和cbaB基因分别编码双加氧酶和还原酶蛋白。将cbaAB基因推导的氨基酸序列与其他加氧酶的序列进行比较，发现在IA类加氧酶中存在一个明确界定的谱系，该谱系具有几个独特特征。这个谱系包括邻苯二甲酸4,5-双加氧酶（pht23），并且根据组分A的可用N端序列，还包括4-磺基苯甲酸3,4-双加氧酶。由vanAB基因编码的香草酸脱甲基酶，正式名称为催化氧化脱甲基的单加氧酶，也包含在这个谱系中。末端氯苯甲酸双加氧酶（CbaA）组分的特征是具有保守的Rieske型[2Fe-2S]R配体中心。还原酶组分（CbaB）包含植物型铁氧还蛋白[2Fe-2S]Fd、FMN-异咯嗪和NAD-核糖结合结构域，并且这些结构域的取向在所有已知的IA类还原酶中都是保守的。这些结果支持这样的假设，即还原酶的电子传递模块的替代融合在加氧酶系统分化的早期就出现了。作用于IA类加氧酶分化的主要进化限制似乎是在芳香环上氧插入位点的对位需要一个羧基。
