A symmetrical tetramer for S. aureus pyruvate carboxylase in complex with coenzyme A.
Pyruvate carboxylase (PC) is a conserved metabolic enzyme with important cellular functions. We report crystallographic and cryo-electron microscopy (EM) studies of Staphylococcus aureus PC (SaPC) in complex with acetyl-CoA, an allosteric activator, and mutagenesis, biochemical, and structural studies of the biotin binding site of its carboxyltransferase (CT) domain. The disease-causing A610T mutation abolishes catalytic activity by blocking biotin binding to the CT active site, and Thr908 might play a catalytic role in the CT reaction. The crystal structure of SaPC in complex with CoA reveals a symmetrical tetramer, with one CoA molecule bound to each monomer, and cryo-EM studies confirm the symmetrical nature of the tetramer. These observations are in sharp contrast to the highly asymmetrical tetramer of Rhizobium etli PC in complex with ethyl-CoA. Our structural information suggests that acetyl-CoA promotes a conformation for the dimer of the biotin carboxylase domain of PC that might be catalytically more competent.
Yu
Linda P C
LP
Department of Biological Sciences, Columbia University, New York, NY 10027, USA.
Xiang
Song
S
Lasso
Gorka
G
Gil
David
D
Valle
Mikel
M
Tong
Liang
L
D000105:Acetyl Coenzyme A / Q000378:metabolism*; D000494:Allosteric Regulation; D000595:Amino Acid Sequence; D001665:Binding Sites; D001710:Biotin / Q000378:metabolism; D019731:Carbon-Nitrogen Ligases / Q000737:chemistry / Q000378:metabolism; D002384:Catalysis; D020285:Cryoelectron Microscopy; D003460:Crystallization; D018360:Crystallography, X-Ray; D019281:Dimerization; D004789:Enzyme Activation; D008958:Models, Molecular; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D011485:Protein Binding; D011487:Protein Conformation; D020836:Protein Structure, Quaternary; D017433:Protein Structure, Secondary; D017434:Protein Structure, Tertiary; D011766:Pyruvate Carboxylase / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism* / Q000648:ultrastructure; D013211:Staphylococcus aureus / Q000201:enzymology*; D014961:X-Ray Diffraction
D001710:Biotin; D000105:Acetyl Coenzyme A; D019731:Carbon-Nitrogen Ligases; C019791:biotin carboxylase; D011766:Pyruvate Carboxylase