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ADP-ribosylation of dinitrogenase reductase from Clostridium pasteurianum prevents its inhibition of nitrogenase from Azotobacter vinelandii.

ABSTRACT: The effect of ADP-ribosylation of dinitrogenase reductase on its binding to dinitrogenase was investigated. Dinitrogenase reductase from Clostridium pasteurianum (Cp2) was a substrate for the ADP-ribosyltransferase and the dinitrogenase-reductase-activating glycohydrolase from Rhodospirillum rubrum. ADP-ribosylation inactivated Cp2 and prevented its formation of a tight complex with dinitrogenase from Azotobacter vinelandii (Av1). The complex between Cp2 and Av1 could not be ADP-ribosylated once it formed.

SUBMITTER: Murrell SA

PROVIDER: S-EPMC1149044 | biostudies-other | 1988 Apr

REPOSITORIES: biostudies-other

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Project description:Biological nitrogen fixation, the microbial reduction of atmospheric nitrogen to bioavailable ammonia, represents both a major limitation on biological productivity and a highly desirable engineering target for synthetic biology. However, the engineering of nitrogen fixation requires an integrated understanding of how the gene regulatory dynamics of host diazotrophs respond across sequence-function space of its central catalytic metalloenzyme, nitrogenase. Here, we interrogate this relationship by analyzing the transcriptome of Azotobacter vinelandii engineered with a phylogenetically inferred ancestral nitrogenase protein variant. The engineered strain exhibits reduced cellular nitrogenase activity but recovers wild-type growth rates following an extended lag period. We find that expression of genes within the immediate nitrogen fixation network is resilient to the introduced nitrogenase sequence-level perturbations. Rather the sustained physiological compatibility with the ancestral nitrogenase variant is accompanied by reduced expression of genes that support trace metal and electron resource allocation to nitrogenase. Our results spotlight gene expression changes in cellular processes adjacent to nitrogen fixation as productive engineering considerations to improve compatibility between remodeled nitrogenase proteins and engineered host diazotrophs. IMPORTANCE Azotobacter vinelandii is a key model bacterium for the study of biological nitrogen fixation, an important metabolic process catalyzed by nitrogenase enzymes. Here, we demonstrate that compatibilities between engineered A. vinelandii strains and nitrogenase variants can be modulated at the regulatory level. The engineered strain studied here responds by adjusting the expression of proteins involved in cellular processes adjacent to nitrogen fixation, rather than that of nitrogenase proteins themselves. These insights can inform future strategies to transfer nitrogenase variants to non-native hosts.

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ADP-ribosylation of dinitrogenase reductase from Clostridium pasteurianum prevents its inhibition of nitrogenase from Azotobacter vinelandii.

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