Crystal structure of the molybdenum cofactor biosynthesis protein MobA from Escherichia coli at near-atomic resolution.
BACKGROUND
All mononuclear molybdoenzymes bind molybdenum in a complex with an organic cofactor termed molybdopterin (MPT). In many bacteria, including Escherichia coli, molybdopterin can be further modified by attachment of a GMP group to the terminal phosphate of molybdopterin to form molybdopterin guanine dinucleotide (MGD). This modification reaction is required for the functioning of many bacterial molybdoenzymes, including the nitrate reductases, dimethylsulfoxide (DMSO) and trimethylamine-N-oxide (TMAO) reductases, and formate dehydrogenases. The GMP attachment step is catalyzed by the cellular enzyme MobA.


RESULTS
The crystal structure of the 21.6 kDa E. coli MobA has been determined by MAD phasing with selenomethionine-substituted protein and subsequently refined at 1. 35 A resolution against native data. The structure consists of a central, predominantly parallel beta sheet sandwiched between two layers of alpha helices and resembles the dinucleotide binding Rossmann fold. One face of the molecule bears a wide depression that is lined by a number of strictly conserved residues, and this feature suggests that this is where substrate binding and catalysis take place.


CONCLUSIONS
Through comparisons with a number of structural homologs, we have assigned plausible functions to several of the residues that line the substrate binding pocket. The enzymatic mechanism probably proceeds via a nucleophilic attack by MPT on the GMP donor, most likely GTP, to produce MGD and pyrophosphate. By analogy with related enzymes, this process is likely to require magnesium ions.
Stevenson
C E
CE
Department of Biological Chemistry and John Innes Centre NR4 7UH, Norwich, United Kingdom.
Sargent
F
F
Buchanan
G
G
Palmer
T
T
Lawson
D M
DM
D000595:Amino Acid Sequence; D001426:Bacterial Proteins / Q000737:chemistry*; D001665:Binding Sites; D020134:Catalytic Domain; D003067:Coenzymes; D016384:Consensus Sequence; D018360:Crystallography, X-Ray; D004926:Escherichia coli / Q000201:enzymology*; D029968:Escherichia coli Proteins; D019143:Evolution, Molecular; D006157:Guanosine Monophosphate / Q000378:metabolism; D008667:Metalloproteins / Q000378:metabolism; D008958:Models, Molecular; D008969:Molecular Sequence Data; D000090006:Molybdenum Cofactors; D011485:Protein Binding; D011487:Protein Conformation; D017433:Protein Structure, Secondary; D011621:Pteridines / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000737:chemistry; D012645:Selenomethionine / Q000737:chemistry; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid
D001426:Bacterial Proteins; D003067:Coenzymes; D029968:Escherichia coli Proteins; D006157:Guanosine Monophosphate; D008667:Metalloproteins; D000090006:Molybdenum Cofactors; D011621:Pteridines; D011993:Recombinant Fusion Proteins; D012645:Selenomethionine; C095754:mobA protein, E coli; C022261:molybdopterin