Isolation, Complementation and Partial Characterization of Mutants of the Methanol Autotroph Xanthobacter H4-14 Defective in Methanol Dissimilation

SUMMARY: Seven mutants of Xanthobacter H4-14, unable to grow on methanol but capable of growth on formate, were isolated and complemented with a chromosomal clone bank constructed in the broad-host-range cosmid pVK100. One mutant could not be complemented but the others fell into four distinct complementation groups that involved three different recombinant clones. All of the complementing regions were separated by at least 10 kbp. The five complementation classes had different phenotypic characteristics and were defective in different aspects of methanol and formaldehyde oxidation. Class I mutants were defective in methanol oxidation, class II mutants were impaired in formaldehyde oxidation, class III mutants appeared to be defective in a regulatory element involving the methanol oxidation system, and class IV mutants appeared to be defective in a regulatory element involving formaldehyde oxidation. Class V mutants exhibited a methanol-sensitive phenotype, which was correlated with an imbalance between methanol and formaldehyde dehydrogenase activities. Analysis of this class suggested it was defective in a repressor that regulated methanol dissimilation functions.