title: Metal chelating properties of adenylate kinase from Paracoccus denitrificans.
abstract: Zinc, a common element of adenylate kinases from Gram-positive bacteria, binds to a structural motif consisting of three or four cysteine residues, Cys-X2-Cys-X16-Cys-X2-Cys/Asp. The enzyme from Paracoccus denitrificans, a Gram-negative bacterium, has structural features much similar to those of adenylate kinases from Gram-positive organisms [Spurgin, P., Tomasselli, A.G., and Schiltz, E. (1989) Eur. J. Biochem., 179, 621-628]. However, adenylate kinase isolated from this bacterium was not reported to bind metal. These findings prompted us to clone the corresponding gene of P. denitrificans, and to characterize the enzyme overproduced in Escherichia coli. The deduced primary structure of adenylate kinase from P. denitrificans revealed two differences from that previously published: Cys was found at position 130 instead of His, and His was found at position 138 instead of Gly. The recombinant enzyme is a dimer which binds either zinc or iron, in a metal/monomer ratio of one. The dissociating sulfhydryl reagent, p-(hydroxy-mercuri)phenylsulfonate, released the metal from the protein, confirming that thiols are involved in zinc- or iron-binding. The iron-chelated form of recombinant P. denitrificans adenylate kinase, which is essentially under reduced form, transfers electrons to the oxidized cytochrome c. In conclusion, the absence of metal in the enzyme isolated from P. denitrificans is not related to the protein structure but most probably due to the physiological properties of the host organism.
authors.affiliation: Laboratoire de Chimie Structurale des Macromolécules, URA CNRS 1129, Institut Pasteur, Paris, France.
authors.forename: V
authors.initials: V
authors.lastname: Perrier
authors.forename: S
authors.initials: S
authors.lastname: Burlacu-Miron
authors.forename: A
authors.initials: A
authors.lastname: Boussac
authors.forename: A
authors.initials: A
authors.lastname: Meier
authors.forename: A M
authors.initials: AM
authors.lastname: Gilles
mesh_terms: D000263:Adenylate Kinase / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D000595:Amino Acid Sequence; D001483:Base Sequence; D002614:Chelating Agents / Q000737:chemistry / Q000378:metabolism*; D003001:Cloning, Molecular; D003545:Cysteine / Q000235:genetics / Q000378:metabolism; D019281:Dimerization; D004228:Dithionitrobenzoic Acid / Q000378:metabolism; D004795:Enzyme Stability; D004926:Escherichia coli / Q000235:genetics; D006639:Histidine / Q000235:genetics / Q000378:metabolism; D007501:Iron / Q000378:metabolism / Q000494:pharmacology; D007700:Kinetics; D008969:Molecular Sequence Data; D010084:Oxidation-Reduction; D010231:Paracoccus denitrificans / Q000201:enzymology* / Q000378:metabolism; D010766:Phosphorylation / Q000187:drug effects; D011489:Protein Denaturation; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000378:metabolism; D017386:Sequence Homology, Amino Acid; D017186:Titrimetry; D014357:Trypsin / Q000378:metabolism; D015032:Zinc / Q000378:metabolism / Q000494:pharmacology
chemical_list: D002614:Chelating Agents; D011994:Recombinant Proteins; D006639:Histidine; D004228:Dithionitrobenzoic Acid; D007501:Iron; D000263:Adenylate Kinase; D014357:Trypsin; D015032:Zinc; D003545:Cysteine