Dipti D. Nayak, Christopher J. Marx

Methylotrophs grow on reduced single carbon compounds like methylamine as the sole source of carbon and energy. In Methylobacterium extorquens AM1, the best-studied aerobic methylotroph, a periplasmic methylamine dehydrogenase that catalyzes the primary oxidation of methylamine to formaldehyde has been examined in great detail. However, recent metagenomic data from natural ecosystems is revealing the abundance and importance of lesser-known routes such as the N-methylglutamate pathway for methylamine oxidation. In this paper we use M. extorquens PA1, a strain closely related to M. extorquens AM1 but lacking the methylamine dehydrogenase, to dissect the genetics and physiology of the ecologically relevant N-methylglutamate pathway for methylamine oxidation. Phenotypic analyses of null mutants in genes encoding enzymes of the N-methylglutamate pathway suggested that γ-glutamylmethylamide synthetase was essential for growth on methylamine as a carbon source but not as a nitrogen source . Furthermore, analysis of M. extorquens PA1 mutants with defects in methylotrophy-specific dissimilatory and assimilatory modules suggested that methylamine use via the N-methylglutamate pathway requires the tetrahydromethanopterin (H4MPT) dependent formaldehyde oxidation pathway but not a complete tetrahydrofolate (H4F) dependent formate assimilation pathway, Additionally, we present genetic evidence that formaldehyde-activating enzyme (FAE) homologs might be involved in methylotrophy. Null mutants of FAE and homologs revealed that FAE and FAE2 influence growth rate and FAE3 influences yield during growth of M. extorquens PA1 on methylamine.