Functional role of the MrpA- and MrpD-homologous protein subunits in enzyme complexes evolutionary related to respiratory chain complex I

Abstract

NADH:quinone oxidoreductase or complex I is a large membrane bound enzyme complex that has evolved from the combination of smaller functional building blocks. Intermediate size enzyme complexes exist in nature that comprise some, but not all of the protein subunits in full size 14-subunit complex I. The membrane spanning complex I subunits NuoL, NuoM and NuoN are homologous to each other and to two proteins from one particular class of Na(+)/H(+) antiporters, denoted MrpA and MrpD. In complex I, these ion transporter protein subunits are prime candidates for harboring important parts of the proton pumping machinery. Using a model system, consisting of Bacillus subtilis MrpA and MrpD deletion strains and a low copy expression plasmid, it was recently demonstrated that NuoN can rescue the strain deleted for MrpD but not that deleted for MrpA, whereas the opposite tendency was seen for NuoL. This demonstrated that the MrpA-type and MrpD-type proteins have unique functional specializations. In this work, the corresponding antiporter-like protein subunits from the smaller enzymes evolutionarily related to complex I were tested in the same model system. The subunits from 11-subunit complex I from Bacillus cereus behaved essentially as those from full size complex I, corroborating that this enzyme should be regarded as a bona fide complex I. The hydrogenase-3 and hydrogenase-4 antiporter-like proteins on the other hand, could substitute equally well for MrpA or MrpD at pH7.4, suggesting that these enzymes have intermediate forms of the antiporter-like proteins, which seemingly lack the functional specificity.

Functional Role of MrpA in the MrpABCDEFG Na + /H + Antiporter Complex from the Archaeon Methanosarcina acetivorans

“…1, supporting the conclusion that MrpA is essential for Na ϩ /H ϩ antiport activity. The results further show that activity is independent of an intact MrpABCDEFG complex, contrary to previous assertions that intact complexes from the domain Bacteria are required for activity (2,3,11,20,25,26).…”

“…Indeed, NuoL and NuoN are functional replacements for MrpA and MrpD of Bacillus species (20). The recently reported crystal structure of complex I highlights the similarity of MrpA and MrpD from the domain Bacteria with antiporter-like subunits NuoL and NuoM/NuoN, respectively, of complex I (21).…”

“…MrpA and MrpD are homologous to the antiporter-like subunits NuoL and NuoM/ NuoN, respectively, of respiratory complex I from the domains Bacteria and Eukarya (18)(19)(20). Indeed, NuoL and NuoN are functional replacements for MrpA and MrpD of Bacillus species (20).…”

“…The seven-subunit MrpABCDEFG complexes from Bacillus species have been model systems for investigation, with a focus on the MrpA and MrpD subunits that contain transmembrane acidic residues, consistent with ion translocation functions (24). Although MrpA and MrpD subunits from the domain Bacteria are implicated in translocation of Na ϩ and H ϩ , respectively, a consensus has emerged that all seven subunits are required for antiport activity (2,3,11,20,25,26). Specifically, expression of an MrpAD subcomplex in the antiporterdeficient Escherichia coli strain KNabc showed no antiport activity (27).…”

New Mode of Energy Metabolism in the Seventh Order of Methanogens as Revealed by Comparative Genome Analysis of “Candidatus Methanoplasma termitum”

“…The C subunit of the ATP synthase (AhaC) has the same conserved Na ϩ -binding motif as in Methanosarcina acetivorans and Methanosarcina mazei, but also in those organisms the ion specificity of ATP synthase is not fully resolved (4). Since all 11-subunit complexes are considered to be proton pumps (61) and since the Fpo-like complex is the only energy-converting complex in Methanomassiliicoccales, it is likely that methanogenesis is coupled to ATP synthesis via a proton motive force.…”