Aerobic soil bacteria metabolize atmospheric hydrogen (H₂) to persist when nutrient sources are limited. This process is the primary sink in the global H₂ cycle and supports the productivity of microbes in oligotrophic environments. To mediate this function, bacteria possess [NiFe]-hydrogenases capable of oxidising H2 to subatmospheric concentrations. The soil saprophyte Mycobacterium smegmatis has two such [NiFe]-hydrogenases, designated Huc and Hhy, which belong to different phylogenetic subgroups. Huc and Hhy exhibit similar characteristics: both are oxygen-tolerant, oxidise H2 to subatmospheric concentrations, and enhance survival during hypoxia and carbon limitation. These shared characteristics pose the question: Why does M. smegmatis require two hydrogenases mediating a seemingly similar function? In this work we resolve this question by showing that Huc and Hhy are differentially expressed, localised, and integrated into the respiratory chain. Huc is active in late exponential and early stationary phase, supporting energy conservation during mixotrophic growth and the transition into dormancy. In contrast, Hhy is most active during long-term persistence, providing energy for maintenance processes when carbon sources are depleted. We show that Huc and Hhy are obligately linked to the aerobic respiratory chain via the menaquinone pool and are differentially affected by respiratory uncouplers. Consistent with their distinct expression profiles, Huc and Hhy interact differentially with the terminal oxidases of the respiratory chain. Huc exclusively donates electrons to, and possibly physically associates with, the proton pumping cytochrome bcc-aa₃ supercomplex. In contrast, the more promiscuous Hhy can also provide electrons to the cytochrome bd oxidase complex. These data demonstrate that, despite their similar characteristics, Huc and Hhy perform distinct functions during mycobacterial growth and survival.

Item Type:	Refereed Article
Keywords:	quinone, mycobacteria, hydrogenase, Mycobacterium smegmatis, respiratory chain
Research Division:	Biological Sciences
Research Group:	Biochemistry and cell biology
Research Field:	Enzymes
Objective Division:	Expanding Knowledge
Objective Group:	Expanding knowledge
Objective Field:	Expanding knowledge in the biological sciences
UTAS Author:	Warr, CG (Professor Coral Warr)
ID Code:	135424
Year Published:	2019
Web of Science® Times Cited:	12
Deposited By:	Medicine
Deposited On:	2019-10-18
Last Modified:	2020-07-28
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