Summary: Two temperature-sensitive mutants of have been isolated and characterized. The first, NH-4, had a temperature-sensitive defect in methylamine oxidation and was unable to utilize methylamine as sole carbon and nitrogen source at 42 °C. The activities of the methylamine oxidation system and methylamine dehydrogenase in cells grown on methylamine at 30 °C were much more thermolabile than those of the wild-type. Furthermore, the affinity of the mutant enzyme for methylamine was lower than that of the wild-type enzyme. These results suggest that NH-4 produces a mutant enzyme with an altered conformation which is more susceptible to thermal denaturation than the wild-type enzyme. Surprisingly, this mutant could grow at 42 °C on media containing methylamine if an alternative carbon or nitrogen source was available. The methylamine oxidation system of whole cells grown under these conditions was not inactivated at 42 °C. The second mutant, NH-7, was unable to grow at the restrictive temperature on media containing either methanol or methylamine as sole carbon source. It contained thermolabile forms of methanol and methylamine dehydrogenases and cytochrome . This phenotype could be due to a mutation in a gene essential for the production of mature forms of these periplasmic proteins, which are involved in C metabolism. Cosmid pAD833, which has previously been shown to carry genes involved in methanol oxidation ( genes), complemented both NH-4 and NH-7. Subcloning indicated that the gene which complemented NH-7 was within a 3 kbp region which contained at least two genes. This region was near an 8 kbp region containing the gene which complemented mutant NH-4.


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