Cloning and expression in Escherichia coli of Acinetobacter calcoaceticus genes for benzoate degradation.
The catabolic genes necessary for the conversion of benzoate to catechol have been cloned from Acinetobacter calcoaceticus into Escherichia coli. The cloned genes, benABCD, encoded both a benzoate 1,2-dioxygenase system, composed of NADH-cytochrome c reductase and terminal oxygenase components, and a cis-diol dehydrogenase. The dioxygenase system appears to be encoded by three genes, benABC, whose products, 53-, 19-, and 38-kilodalton proteins, correspond in size to those of components in other bacterial dioxygenases. The cloned dioxygenase system is expressed at high level in E. coli, enabling the conversion of benzoate to a cis-diol, 2-hydro-1,2-dihydroxybenzoate, at a rate comparable to that of fully induced A. calcoaceticus cultures. A cis-diol dehydrogenase, the product of the A. calcoaceticus benD gene, when present in E. coli enables this organism to convert the cis-diol intermediate to catechol. The dehydrogenase has been partially purified and is a dimer with two identical 31-kilodalton subunits. The ben genes are clustered on the A. calcoaceticus chromosome with independently regulated genes needed for the dissimilation of catechol. In a 16-kilobase-pair region of the chromosome there are 10 genes for benzoate catabolism, organized in no fewer than three transcriptional units. This kind of arrangement, termed supraoperonic clustering, has been observed previously in pseudomonads.
Neidle
E L
EL
Department of Biology, Yale University, New Haven, Connecticut 06511.
Shapiro
M K
MK
Ornston
L N
LN
D000150:Acinetobacter / Q000201:enzymology / Q000235:genetics* / Q000378:metabolism; D001565:Benzoates / Q000378:metabolism*; D001673:Biodegradation, Environmental; D002396:Catechols / Q000378:metabolism; D055598:Chemical Phenomena; D002621:Chemistry; D003001:Cloning, Molecular; D004262:DNA Restriction Enzymes; D004269:DNA, Bacterial / Q000235:genetics; D004926:Escherichia coli / Q000235:genetics*; D005786:Gene Expression Regulation; D005798:Genes, Bacterial; D009154:Mutation; D010088:Oxidoreductases / Q000235:genetics; D010105:Oxygenases / Q000235:genetics; D014169:Transformation, Bacterial
D001565:Benzoates; D002396:Catechols; D004269:DNA, Bacterial; D010088:Oxidoreductases; D010105:Oxygenases; C019766:benzoate 1,2-dioxygenase; C025312:3,5-cyclohexadiene-1,2-diol-1-carboxylate dehydrogenase; D004262:DNA Restriction Enzymes