Involvement of a mate chaperone (TorD) in the maturation pathway of molybdoenzyme TorA.
As many prokaryotic molybdoenzymes, the trimethylamine oxide reductase (TorA) of Escherichia coli requires the insertion of a bis(molybdopterin guanine dinucleotide)molybdenum cofactor in its catalytic site to be active and translocated to the periplasm. We show in vitro that the purified apo form of TorA was activated weakly when an appropriate bis(molybdopterin guanine dinucleotide)molybdenum source was provided, whereas addition of the TorD chaperone increased apoTorA activation up to 4-fold, allowing maturation of most of the apoprotein. We demonstrate that TorD alone is sufficient for the efficient activation of apoTorA by performing a minimal in vitro assay containing only the components for the cofactor synthesis, apoTorA and TorD. Interestingly, incubation of apoTorA with TorD before cofactor addition led to a significant increase of apoTorA activation, suggesting that TorD acts on apoTorA before cofactor insertion. This result is consistent with the fact that TorD binds to apoTorA and probably modifies its conformation in the absence of cofactor. Therefore, we propose that TorD is involved in the first step of TorA maturation to make it competent to receive the cofactor.
Ilbert
Marianne
M
Laboratoire de Chimie Bactérienne and Laboratoire de Bioénergétique et Ingénierie des Protéines, Institut de Biologie Structurale et Microbiologie, CNRS, 31, chemin Joseph Aiguier, 13402 Marseille Cedex 20, France.
Méjean
Vincent
V
Giudici-Orticoni
Marie-Thérèse
MT
Samama
Jean-Pierre
JP
Iobbi-Nivol
Chantal
C
D001059:Apoproteins / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D004591:Electrophoresis, Polyacrylamide Gel; D004789:Enzyme Activation / Q000187:drug effects; D004926:Escherichia coli / Q000201:enzymology*; D029968:Escherichia coli Proteins / Q000302:isolation & purification / Q000494:pharmacology*; D006150:Guanine Nucleotides / Q000494:pharmacology; D007700:Kinetics; D018832:Molecular Chaperones / Q000302:isolation & purification / Q000494:pharmacology*; D010089:Oxidoreductases, N-Demethylating / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism*; D011487:Protein Conformation / Q000187:drug effects; D011622:Pterins / Q000494:pharmacology
D001059:Apoproteins; D029968:Escherichia coli Proteins; D006150:Guanine Nucleotides; D018832:Molecular Chaperones; D011622:Pterins; C088083:TorD protein, E coli; C063761:molybdopterin guanine dinucleotide; D010089:Oxidoreductases, N-Demethylating; C023186:trimethylamine dehydrogenase