Development of a Rhodobacter capsulatus self‐reporting model system for optimizing light‐dependent, [FeFe]‐hydrogenase‐driven H₂ production

Abstract

The photosynthetic bacterium Rhodobacter capsulatus normally photoproduces H as a by-product of its nitrogenase-catalyzed nitrogen-fixing activity. Such H production, however, is expensive from a metabolic perspective, requiring nearly four times as many photons as the equivalent algal hydrogenase-based system (Ghirardi et al., 2009 Photobiological hydrogen-producing systems. Chem Soc Rev 38(1):52-61). Here, we report the insertion of a Clostridium acetobutylicum [FeFe]-hydrogenase and its three attendant hydrogenase assembly proteins into an R. capsulatus strain lacking its native uptake hydrogenase. Further, this strain is modified to fluoresce upon sensing H . The resulting strain photoproduces H and self-reports its own H production through fluorescence. This model system represents a unique method of developing hydrogenase-based H production in R. capsulatus, may serve as a powerful system for in vivo directed evolution of hydrogenases and hydrogenase-associated genes, and provides a means of screening for increased metabolic production of H . Biotechnol. Bioeng. 2017;114: 291-297. © 2016 Wiley Periodicals, Inc.

Toward a synthetic hydrogen sensor in cyanobacteria: Functional production of an oxygen-tolerant regulatory hydrogenase in Synechocystis sp. PCC 6803

“…We successfully implemented two parts of the four-part H 2 -responsive signal transduction cascade from C. necator in the cyanobacterium Synechocystis. This is considered as a first step toward a synthetic cyanobacterial H 2 biosensor that could be used analogously to previous reports (Wecker et al, 2017). A potential application would be, for instance, to optimize the H 2 evolution activities of heterologously produced O 2 -tolerant, energy-converting hydrogenases, one of which was successfully implemented in Synechocystis recently (Lupacchini et al, 2021).…”

“…For example, Wecker et al used engineered R. capsulatus strains that monitor H 2 by reporter fluorescence to track the activity of a recombinant H 2 -evolving hydrogenase from Clostridium acetobutylicum . Only low H 2 production has been detected, but the system potentially enables further screening and hydrogenase evolution approaches ( Wecker et al, 2017 ). We successfully implemented two parts of the four-part H 2 -responsive signal transduction cascade from C. necator in the cyanobacterium Synechocystis .…”

Development of novel strategies for higher fermentative biohydrogen recovery along with novel metabolites from organic wastes: The present state of the art

“…Wecker et al. [90] reported hup ‐based bioH 2 production by R. capsulatus cells. This strain was initially deprived of native uptake hydrogenase.…”

“…Elimination of hup from the genome of R. sphaeroides cells enhanced bioH 2 production rates by 11-40% as compared to the wild strain. Wecker et al [90] reported hup-based bioH 2 production by R. capsulatus cells. This strain was initially deprived of native uptake hydrogenase.…”