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2‐Oxoglutarate and the PII homologues NifI 1 and NifI 2 regulate nitrogenase activity in cell extracts of Methanococcus maripaludis
Author(s) -
Dodsworth Jeremy A.,
Cady Nathaniel C.,
Leigh John A.
Publication year - 2005
Publication title -
molecular microbiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.857
H-Index - 247
eISSN - 1365-2958
pISSN - 0950-382X
DOI - 10.1111/j.1365-2958.2005.04621.x
Subject(s) - nitrogenase , biology , mutant , methanococcus , biochemistry , wild type , enzyme , nitrogen fixation , bacteria , gene , archaea , genetics
Summary Post‐translational regulation of nitrogen fixation, or switch‐off, in the methanogenic archaeon Methanococcus maripaludis does not involve detectable covalent modification of the dinitrogenase reductase as in some bacteria, and the genes encoding the PII homologues NifI 1 and NifI 2 are both required, indicating a novel mechanism. To further understand the mechanism of switch‐off, we assayed nitrogenase activity in cell extracts from wild‐type and nifI mutant strains in the absence or presence of potential signals of nitrogen status. Activity in extracts from a Δ nifI 1 nifI 2 strain was sixfold higher than in extracts from wild‐type cells. Addition of 2‐oxoglutarate to wild‐type extracts enhanced activity up to fivefold, a level similar to that observed in Δ nifI 1 nifI 2 extracts. 2‐Oxoglutarate did not affect activity in Δ nifI 1 nifI 2 or single nifI mutant extracts. Furthermore, extracts from genetically complimented nifI mutants regained wild‐type characteristics, indicating an in vitro correlation with in vivo effects. Extraction and quantification of 2‐oxoglutarate indicated concentrations 10‐fold higher in nitrogen‐fixing cells than in switched‐off and ammonium‐grown cells. We propose a model for switch‐off where the NifI proteins have an inhibitory effect on nitrogenase activity that is counteracted by high levels of 2‐oxoglutarate, which acts as a signal of nitrogen limitation.