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Organization and control of genes encoding catabolic enzymes in Rhizobiaceae. Progress report, March 1993
Rhizobiaceae, a diverse bacterial group comprising rhizobia and agrobacteria, symbiotic partnership with plants form nitrogen-fixing nodules on plant roots or are plant pathogens. Phenolic compounds produced by plants serve as inducers of rhizobial nodulation genes and agrobacterial virulence genes reflect their capacity to utilize numerous aromatics, including phenolics, as a source of carbon and energy. In many microbes the aerobic degradation of numerous aromatic compounds to tricarboxylic acid cycle intermediates is achieved by the {beta}-ketoadipate pathway. Our initial studies focused on the organization and regulation of the ketoadipate pathway in Agrobacterium tumefaciens. We have cloned, identified and characterized a novel regulatory gene that modulates expression of an adjacent pca (protocatechuate) structural gene, pcaD. Regulation of pcaD is mediated by the regulatory gene, termed pcaQ, in concert with the intermediate {beta}-carboxy-cis,cis-muconate. {beta}-carboxy-cis,cismuconate is an unstable chemical, not marketed commercially, and it is unlikely to permeate Escherichia coli cells if supplied in media. Because of these factors, characterization of pcaQ in E. coli required an in vivo delivery system for {beta}-carboxycis,cis-muconate. This was accomplished by designing an E. coli strain that expressed an Acinetobacter calcoaceticus pcaA gene for conversion of protocatechuate to {beta}-carboxy-cis,cis-muconate
Abundant expression of Pseudomonas genes for chlorocatechol metabolism.
The respective specific activities of catechol 1,2-oxygenase II (catechol 1,2-dioxygenase; EC 1.13.11.1) and muconate cycloisomerase II (chloromuconate cycloisomerase; EC 5.5.1.7) in crude extracts of chlorobenzoate-grown Pseudomonas cells corresponded to about 16 and 11% of the soluble cell protein. High levels of protein synthesis appeared to compensate for a loss in catalytic activity that accompanied evolutionary acquisition of broad substrate specificity required for the enzymes to accommodate halogenated substrates
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