Crystal structure of pyruvate kinase from Geobacillus stearothermophilus.
The pyruvate kinase (PK) from a moderate thermophile, Geobacillus stearothermophilus, is an allosteric enzyme activated by AMP and ribose 5-phosphate but not fructose 1, 6-bisphosphate (FBP), which is a common activator of PKs. It has an extra C-terminal sequence (ECTS), which contains a highly conserved phosphoenolpyruvate (PEP) binding motif, but its function and structure remain unclear. To elucidate the structural characteristics of the effector-binding site and the ECTS, the crystal structure of the C9S/C268S mutant of the enzyme was determined at 2.4 A resolution. The crystal belonged to space group P6(2)22, with unit cell parameters a, b = 145.97 A, c = 118.03 A. The enzyme was a homotetramer and its overall domain structure was similar to the previously solved structures except that the ECTS formed a new domain (C' domain). The structure of the C' domain closely resembled that of the PEP binding domain of maize pyruvate phosphate dikinase. A sulphate ion was found in a pocket in the effector-binding C domain. This site corresponds to the 6-phosphate group-binding site in yeast PK bound FBP and seems to be the effector-binding site. Through comparison of the structure of the putative effector-binding site to that of the FBP binding site of the yeast enzyme, the structural basis of the effector specificity of the G. stearothermophilus PK is discussed.
Suzuki
Kenichiro
K
Department of Food and Nutritional Sciences, University of Shizuoka, Yada 52-1, Shizuoka 422-8526, Japan.
Ito
Sohei
S
Shimizu-Ibuka
Akiko
A
Sakai
Hiroshi
H
D000249:Adenosine Monophosphate / Q000737:chemistry; D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D018360:Crystallography, X-Ray; D001411:Geobacillus stearothermophilus / Q000201:enzymology*; D006801:Humans; D007700:Kinetics; D008969:Molecular Sequence Data; D009154:Mutation; D011487:Protein Conformation; D017434:Protein Structure, Tertiary; D011770:Pyruvate Kinase / Q000737:chemistry*; D012267:Ribosemonophosphates / Q000737:chemistry; D017386:Sequence Homology, Amino Acid; D003313:Zea mays / Q000201:enzymology
D012267:Ribosemonophosphates; D000249:Adenosine Monophosphate; C031626:ribose-5-phosphate; D011770:Pyruvate Kinase