NarJ chaperone binds on two distinct sites of the aponitrate reductase of Escherichia coli to coordinate molybdenum cofactor insertion and assembly.
Understanding when and how metal cofactor insertion occurs into a multisubunit metalloenzyme is of fundamental importance. Molybdenum cofactor insertion is a tightly controlled process that involves specific interactions between the proteins that promote cofactor delivery, enzyme-specific chaperones, and the apoenzyme. In the assembly pathway of the multisubunit molybdoenzyme, membrane-bound nitrate reductase A from Escherichia coli, a NarJ-assisted molybdenum cofactor (Moco) insertion step, must precede membrane anchoring of the apoenzyme. Here, we have shown that the NarJ chaperone interacts at two distinct binding sites of the apoenzyme, one interfering with its membrane anchoring and another one being involved in molybdenum cofactor insertion. The presence of the two NarJ-binding sites within NarG is required to ensure productive formation of active nitrate reductase. Our findings supported the view that enzyme-specific chaperones play a central role in the biogenesis of multisubunit molybdoenzymes by coordinating subunits assembly and molybdenum cofactor insertion.
Vergnes
Alexandra
A
Laboratoire de Chimie Bactérienne, Institut Biologie Structurale et Microbiologie (IBSM), Centre National de la Recherche Scientifique, 31 chemin Joseph Aiguier, 13402 Marseille cedex 09, France.
Pommier
Janine
J
Toci
René
R
Blasco
Francis
F
Giordano
Gérard
G
Magalon
Axel
A
D001665:Binding Sites; D015374:Biosensing Techniques; D004926:Escherichia coli / Q000201:enzymology* / Q000378:metabolism; D029968:Escherichia coli Proteins / Q000737:chemistry*; D017353:Gene Deletion; D008954:Models, Biological; D008958:Models, Molecular; D018832:Molecular Chaperones / Q000737:chemistry*; D008982:Molybdenum / Q000737:chemistry*; D050901:Nitrate Reductase / Q000737:chemistry*; D009566:Nitrates / Q000737:chemistry*; D010957:Plasmids / Q000378:metabolism; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D013347:Subcellular Fractions; D013824:Thermus / Q000201:enzymology; D020798:Two-Hybrid System Techniques
D029968:Escherichia coli Proteins; D018832:Molecular Chaperones; C508364:NarJ protein, E coli; D009566:Nitrates; D008982:Molybdenum; C508365:NarG protein, E coli; D050901:Nitrate Reductase