Kinetic and Structural Characterization of Phosphofructokinase from Lactobacillus bulgaricus

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• 作者：N. Monique Paricharttanakul ; Sheng Ye ; Ann L. Menefee ; Farah Javid-Majd ; James C. Sacchettini ; and Gregory D. Reinhart
• 刊名：Biochemistry
• 出版年：2005
• 出版时间：November 22, 2005
• 年：2005
• 卷：44
• 期：46
• 页码：15280 - 15286
• 全文大小：298K
• 年卷期：v.44,no.46(November 22, 2005)
• ISSN：1520-4995

文摘

Phosphofructokinase from Lactobacillus delbrueckii subspecies bulgaricus (LbPFK) has beenreported to be a nonallosteric analogue of phosphofructokinase from Escherichia coli at pH 8.2 [Le Braset al. (1991) Eur. J. Biochem. 198, 683-687]. A reexamination of the kinetics of this enzyme showsLbPFK to have limited binding affinity toward the allosteric ligands, MgADP and PEP, with dissociationconstants of approximately 20 mM for both. Their allosteric effects are observed only at high concentrationsof these ligands, with both exhibiting inhibitory effects on substrate binding. No pH dependence wasobserved for the binding and the influence of MgADP and PEP on the enzyme. To attempt to explainthese results, the crystal structure of LbPFK was solved using molecular replacement to 1.86 Å resolution.A comparative study of the LbPFK structure with that of phosphofructokinases from E. coli (EcPFK) andBacillus stearothermophilus (BsPFK) reveals a structure with conserved fold and substrate binding site.The effector binding site, however, shows many differences that could explain the observed decreases inbinding affinity for MgADP and PEP in LbPFK as compared to the other two enzymes.