Abstract
Carbon monoxide (CO), well known as a toxic gas, is increasingly recognized as a key metabolite and signaling molecule. Microbial utilization of CO is quite common, evidenced by the rapid escalation in description of new species of CO-utilizing bacteria and archaea. Carbon monoxide dehydrogenase (CODH), the protein complex that enables anaerobic CO-utilization, has been well-characterized from an increasing number of microorganisms, however the regulation of multiple CO-related gene clusters in single isolates remains unexplored. Many species are extraordinarily resistant to high CO concentrations, thriving under pure CO at more than one atmosphere. We hypothesized that, in strains that can grow exclusively on CO, both carbon acquisition via the CODH/acetyl CoA synthase complex and energy conservation via a CODH-linked hydrogenase must be differentially regulated in response to the availability of CO. The CO-sensing transcriptional activator, CooA is present in most CO-oxidizing bacteria. Here we present a genomic and phylogenetic survey of CODH operons and cooA genes found in CooA-containing bacteria. Two distinct groups of CooA homologs were found: one clade (CooA-1) is found in the majority of CooA-containing bacteria, whereas the other clade (CooA-2) is found only in genomes that encode multiple CODH clusters, suggesting that the CooA-2 might be important for cross-regulation of competing CODH operons. Recombinant CooA-1 and CooA-2 regulators from the prototypical CO-utilizing bacterium Carboxydothermus hydrogenoformans were purified, and promoter binding analyses revealed that CooA-1 specifically regulates the hydrogenase-linked CODH, whereas CooA-2 is able to regulate both the hydrogenase-linked CODH and the CODH/ACS operons. These studies point to the ability of dual CooA homologs to partition CO into divergent CO-utilizing pathways resulting in efficient consumption of a single limiting growth substrate available across a wide range of concentrations.
Highlights
Anaerobic microbial Carbon monoxide (CO)-utilization, which was first characterized by Uffen (1976) in a mesophilic α-proteobacterium, Rhodopseudomonas spp., is recognized as a widespread metabolic capacity (King and Weber, 2007; Techtmann et al, 2009)
The CooA-2 genes found to date are all found in the genomes of Firmicute bacteria that encode multiple Carbon monoxide dehydrogenase (CODH) operons
C. hydrogenoformans was chosen as a model system to explore the role of CooA-2 in regulating the CODH/ACS since it contains all the genetic elements in the CO “toolkit.”
Summary
Anaerobic microbial CO-utilization, which was first characterized by Uffen (1976) in a mesophilic α-proteobacterium, Rhodopseudomonas spp., is recognized as a widespread metabolic capacity (King and Weber, 2007; Techtmann et al, 2009). Carbon monoxide is ubiquitous in geothermal environments and surprisingly common in aerobic microbial ecosystems, albeit in low concentrations (Dunfield and King, 2005; Techtmann et al, 2009) In both aerobic and anaerobic species, optimal conditions for growth span a wide range of CO concentrations. Hydrogenogenic carboxydotrophs utilize the electrons generated from the oxidation of CO for generation of hydrogen via a membrane bound hydrogenase This physiology is based on the water gas shift reaction CO + H2O → CO2 + H2 (Svetlitchnyi et al, 1991; Kerby et al, 1992, 1995; Sokolova et al, 2004b). Other species shunt the electrons generated from CO oxidation to the reduction of various metals, such as iron or manganese (Slobodkin et al, 1997; Sokolova et al, 2004a)
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