Ring-opening mechanism revealed by crystal structures of NagB and its ES intermediate complex.
Glucosamine 6-phosphate deaminase (NagB) catalyzes the conversion of d-glucosamine 6-phosphate (GlcN6P) to d-fructose 6-phosphate and ammonia. This reaction is the final step of N-acetylglucosamine utilization and decides its metabolic fate. The enzyme from Streptococcus mutans belongs to the monomeric subfamily of NagB. The crystal structure of the native SmuNagB (NagB from S. mutans) presented here, compared with the structures of its homologs BsuNagB (NagB from Bacillus subtilis) and EcoNagB (NagB from E. coli), implies a conformational change of the 'lid' motif in the activation of the monomeric NagB enzyme. We have also captured the enzyme-substrate intermediate complex of the NagB family at low pH, where a remarkable loss of the catalytic activity of SmuNagB was detected. The enzyme-substrate intermediate presents the initial step of the GlcN6P deaminase reaction. The structural evidence (1) supports the alpha-anomer of GlcN6P as the specific natural substrate of NagB; (2) displays the substrate-binding pocket at the active site; and (3) together with the site-directed mutagenesis studies, demonstrates the ring-opening mechanism of an Asn-His-Glu triad that performs the proton transfer from O1 to O5 to open the sugar ring.
Liu
Cong
C
National Laboratory of Protein Engineering and Plant Genetic Engineering, College of Life Sciences, Peking University, Beijing 100871, China.
Li
Dan
D
Liang
Yu-He
YH
Li
Lan-Fen
LF
Su
Xiao-Dong
XD
D019747:Aldose-Ketose Isomerases / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D001412:Bacillus subtilis / Q000201:enzymology; D001426:Bacterial Proteins / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D002384:Catalysis; D020134:Catalytic Domain; D018360:Crystallography, X-Ray; D004926:Escherichia coli / Q000201:enzymology; D005944:Glucosamine / Q000031:analogs & derivatives* / Q000737:chemistry / Q000378:metabolism; D019298:Glucose-6-Phosphate / Q000031:analogs & derivatives* / Q000737:chemistry / Q000378:metabolism; D006863:Hydrogen-Ion Concentration; D008969:Molecular Sequence Data; D011487:Protein Conformation; D013295:Streptococcus mutans / Q000201:enzymology*
D001426:Bacterial Proteins; C001293:glucosamine 6-phosphate; D019298:Glucose-6-Phosphate; C020582:glucosamine-6-phosphate isomerase; D019747:Aldose-Ketose Isomerases; D005944:Glucosamine