title: Mechanism of proton transfer inhibition by Cd(2+) binding to bacterial reaction centers: determination of the pK(A) of functionally important histidine residues.
abstract: The bacterial photosynthetic reaction center (RC) uses light energy to catalyze the reduction of a bound quinone molecule Q(B) to quinol Q(B)H(2). In RCs from Rhodobacter sphaeroides the protons involved in this process come from the cytoplasm and travel through pathways that involve His-H126 and His-H128 located near the proton entry point. In this study, we measured the pH dependence from 4.5 to 8.5 of the binding of the proton transfer inhibitor Cd(2+), which ligates to these surface His in the RC and inhibits proton-coupled electron transfer. At pH <6, the negative slope of the logarithm of the dissociation constant, K(D), versus pH approaches 2, indicating that, upon binding of Cd(2+), two protons are displaced; i.e., the binding is electrostatically compensated. At pH >7, K(D) becomes essentially independent of pH. A theoretical fit to the data over the entire pH range required two protons with pK(A) values of 6.8 and 6.3 (+/-0.5). To assess the contribution of His-H126 and His-H128 to the observed pH dependence, K(D) was measured in mutant RCs that lack the imidazole group of His-H126 or His-H128 (His --> Ala). In both mutant RCs, K(D) was approximately pH independent, showing that Cd(2+) does not displace protons upon binding in the mutant RCs, in contrast to the native RC in which His-H126 and His-H128 are the predominant contributors to the observed pH dependence of K(D). Thus, Cd(2+) inhibits RC function by binding to functionally important histidines.
authors.affiliation: Department of Physics 0319, University of California at San Diego, 9500 Gilman Drive, La Jolla, California 92093, USA.
authors.forename: M L
authors.initials: ML
authors.lastname: Paddock
authors.forename: L
authors.initials: L
authors.lastname: Sagle
authors.forename: A
authors.initials: A
authors.lastname: Tehrani
authors.forename: J T
authors.initials: JT
authors.lastname: Beatty
authors.forename: G
authors.initials: G
authors.lastname: Feher
authors.forename: M Y
authors.initials: MY
authors.lastname: Okamura
mesh_terms: D001224:Aspartic Acid / Q000737:chemistry / Q000378:metabolism; D001665:Binding Sites; D002104:Cadmium / Q000737:chemistry / Q000378:metabolism* / Q000494:pharmacology*; D004579:Electron Transport; D006639:Histidine / Q000737:chemistry* / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008958:Models, Molecular; D016297:Mutagenesis, Site-Directed; D045322:Photosynthetic Reaction Center Complex Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism*; D011485:Protein Binding; D011522:Protons; D011809:Quinones / Q000737:chemistry / Q000378:metabolism; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012242:Rhodobacter sphaeroides / Q000378:metabolism
chemical_list: D045322:Photosynthetic Reaction Center Complex Proteins; D011522:Protons; D011809:Quinones; D011994:Recombinant Proteins; D002104:Cadmium; D001224:Aspartic Acid; D006639:Histidine