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Structure of IDP00960

1.9 Angstrom resolution crystal structure of a NAD synthetase (nadE) from Salmonella typhimurium LT2 in complex with NAD(+)

Edit deposit information
CSGID target
IDP00960 
PDB Id
3HMQ (NCBI MMDB
Authors
A.S.Halavaty,Z.Wawrzak,T.Skarina,O.Onopriyenko,S.N.Peterson,A.Savchenko,W.F.Anderson,Center For Structural Genomics Of Infectious Diseases (Csgid) 
Responsible person
Andrei Halavaty 
Responsible lab
Northwestern University 
Deposition Date
May 29, 2009 
Release Date
Jun 16, 2009 

Annotation

Description
Nicotinamide adenine dinucleotide (NAD) is a ubiquitous carrier of reduction equivalents. It functions as a cofactor in numerous metabolic reactions and is essential for calcium mobilization, DNA repair, and post-translational modification of proteins in eukaryotes. The final step of the NAD biosynthesis is a two-step reaction, which is catalyzed by the NAD synthetases (NADS). Initially, the enzymes mediate an adenylyl transfer from adenosine triphosphate (ATP) to the nicotinysyl moiety of nicotinic acid adenine dinucleotide (NAAD), resulting in NAD-adenylate as an intermediate. Finally, the amidation of the nicotinysyl moiety using either ammonia or glutamine as an amide source results in producing of NAD. The crystal structure of the ammonia-dependent homodimeric NADS from S. typhimurium LT2 in complex with the reaction product NAD is reported. Pairwise structural alignment reveals similar tertiary and quaternary architecture between the NadE protein and other known NH3-dependent NADS structures. This suggests a resembling molecular mechanism of the catalyzed reaction. The 3D structure of the enzyme disclosed two NAAD/NAD binding sites at the dimer interface and empty adenosine triphosphate (ATP) binding site within each subunit. Several sulfate ions have been modeled with two anions docking similar to two phosphates of PPi in the NADS structure from B. subtilis (PDB ID 1EE1). Distortion of the region encompassing residues 209 through 221 of the S. typhimurium enzyme has functional character. In the NAAD-ATP-bound structure of the B. subtilis protein similar region exhibits a loop conformation accommodating two substrates. In the absence of ATP, but presence of NAAD as in the E.coli NAAD-complexed synthetase structure (PDB ID 1WXG), this loops is disordered. That is, adopting stable conformation by this area requires binding of both substrates fulfilling the recognition role of the loop. Association of Mg2+ in the active site is crucial for initiation of the reaction forming the intermediate, which is then cleaved by ammonia releasing all the products and leading to unfolding of the loop region.  
Functional assignment
NAD synthetase 

Ligands

Ligand code Name Ligand type
SO4 crystallization
MSE modified residue
175 3,5-dihydro-5-methylidene-4h-imidazol-4-on biological

Structure information

Unit cell parameters

Space Group
P 31 2 1  
Unit Cell

a=91.82Å, b=91.82Å, c=75.05Å
α=90.00, β=90.00, γ=120.00 
Solvent content
58.64  
Matthews coefficient
2.97  

Refinement

Data for the highest resolution shell is in parentheses.
Resolution range
29.06-1.90Å (1.95-1.90Å)  
Rall(%)
14.6 
Rwork(%)
14.4 (17.4) 
Rfree(%)
18.7 (19.8) 
Num. observed reflections
29124 (2142) 
Num. Rfree reflections
1485 (98) 
Completeness(%)
99.7 (100.0) 

Model parameters

Num Atoms
2161  
Num Waters
432  
Num Hetatoms
557  
Model mean isotropic B factor
19.530Å2  
RMSD bond length
0.011Å  
RMSD bond angle
1.374°  
Filename uploaded
3HMQ.pdb (uploaded on Sep 17, 2010 6:35 PM)  
Inserted
Jun 02, 2009