title: Bicarbonate binding activity of the CmpA protein of the cyanobacterium Synechococcus sp. strain PCC 7942 involved in active transport of bicarbonate.
abstract: The cmpABCD operon of the cyanobacterium Synechococcus sp. strain PCC 7942 encodes an ATP-binding cassette transporter involved in HCO(3)(-) uptake. The three genes, cmpBCD, encode membrane components of an ATP-binding cassette transporter, whereas cmpA encodes a 42-kDa cytoplasmic membrane protein, which is 46.5% identical to the membrane-anchored substrate-binding protein of the nitrate/nitrite transporter. Equilibrium dialysis analysis using H(14)CO(3)(-) showed that a truncated CmpA protein lacking the N-terminal 31 amino acids, expressed in Escherichia coli cells as a histidine-tagged soluble protein, specifically binds inorganic carbon (CO(2) or HCO(3)(-)). The addition of the recombinant CmpA protein to a buffer caused a decrease in the concentration of dissolved CO(2) because of the binding of inorganic carbon to the protein. The decrease in CO(2) concentration was accelerated by the addition of carbonic anhydrase, indicating that HCO(3)(-), but not CO(2), binds to the protein. Mass spectrometric measurements of the amounts of unbound and bound HCO(3)(-) in CmpA solutions containing low concentrations of inorganic carbon revealed that CmpA binds HCO(3)(-) with high affinity (K(d) = 5 microm). A similar dissociation constant was obtained by analysis of the competitive inhibition of the CmpA protein on the carboxylation of phosphoenolpyruvate by phosphoenolpyruvate carboxylase at limiting concentrations of HCO(3)(-). These findings showed that the cmpA gene encodes the substrate-binding protein of the HCO(3)(-) transporter.
authors.affiliation: Molecular Plant Physiology Group, Research School of Biological Sciences, Australian National University, P.O. Box 475, Canberra ACT 2601, Australia.
authors.forename: S
authors.initials: S
authors.lastname: Maeda
authors.forename: G D
authors.initials: GD
authors.lastname: Price
authors.forename: M R
authors.initials: MR
authors.lastname: Badger
authors.forename: C
authors.initials: C
authors.lastname: Enomoto
authors.forename: T
authors.initials: T
authors.lastname: Omata
mesh_terms: D018528:ATP-Binding Cassette Transporters / Q000235:genetics / Q000378:metabolism*; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism*; D001639:Bicarbonates / Q000378:metabolism*; D002245:Carbon Dioxide / Q000378:metabolism; D002256:Carbonic Anhydrases / Q000378:metabolism; D000458:Cyanobacteria / Q000378:metabolism*; D004926:Escherichia coli; D007700:Kinetics; D013058:Mass Spectrometry; D009154:Mutation; D010730:Phosphoenolpyruvate Carboxylase / Q000037:antagonists & inhibitors; D011485:Protein Binding; D011994:Recombinant Proteins / Q000378:metabolism
chemical_list: D018528:ATP-Binding Cassette Transporters; D001426:Bacterial Proteins; D001639:Bicarbonates; D011994:Recombinant Proteins; D002245:Carbon Dioxide; D010730:Phosphoenolpyruvate Carboxylase; D002256:Carbonic Anhydrases