Recent Papers

Williams P.A. & Shaw L.M. (1997) mucK, a gene in Acinetobacter calcoaceticus ADP1 (BD413) encodes the ability to grow on exogeneous cis,cis-muconate as the sole carbon source. J. Bacteriol.179:5935-5942.

ABSTRACT

Benzyl alcohol, benzaldehyde, benzoate and anthranilate are metabolised via catechol, cis,cis-muconate and the ß-ketoadipate pathway in Acinetobacter calcoaceticus ADP1 (BD413). Mutant strain ISA25 with a deletion spanning catBCIJF and unable to metabolise muconate further will not grow in the presence of an aromatic precursor of muconate. Growth on fumarate as sole carbon source with added benzyl alcohol or benzaldehyde selected spontaneous mutants of ISA25. After repair of the cat deletion by natural transformation with linearised plasmid pPAN4 (catBCIJF ) 10 mutants were unable to grow on benzoate or cis,cis-muconate but could still grow on anthranilate. Transformation with wild type chromosomal DNA demonstrated the presence of two unlinked mutations in each strain, one in the benABCD region, encoding the conversion of benzoate to catechol, and the other in a gene determining the ability to grow on exogeneous cis,cis-muconate. The wild-type gene, named mucK, was cloned into pUC18 and its nucleotide sequence determined. It encodes a 413 residue protein of Mr=45252 which is a member of a superfamily of membrane transport proteins and within a subgroup involved in the uptake of organic acids. Five of the mutant alleles were cloned and the mutations determined by nucleotide sequencing. All the mutations were in the mucK coding region and comprised three deletions and one duplication and a substitution. Insertional inactivation of mucK resulted in loss of the ability to utilise exogeneous muconate. Linkage between mucK and pcaU showed that its location on the chromosome appeared to be unique for genes associated with the benzoate branch of the ß-ketoadipate pathway in being close to the pca-qui-pob gene cluster (for p-hydroxybenzoate utilisation) and distant from the functionally related ben-cat cluster. Downstream of mucK and transcribed in the same direction is an ORF of 570 residues (Mr=63002) which shows considerable homology with a mammalian electron transport protein; its insertional inactivation had no detectable phenotypic effect.