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pH Dependence of Catalysis by Pseudomonas aeruginosa Isochorismate-Pyruvate Lyase: Implications for Transition State Stabilization and the Role of Lysine 42
Olucha, José Ouellette, Andrew Nicholas Luo, Qianyi Lamb, Audrey L.
Olucha, José
Ouellette, Andrew Nicholas
Luo, Qianyi
Lamb, Audrey L.
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Abstract
An isochorismate-pyruvate lyase with adventitious chorismate mutase activity from Pseudomonas aerugionsa (PchB) achieves catalysis of both pericyclic reactions in part by the stabilization of reactive conformations and in part by electrostatic transition-state stabilization. When the active site loop Lys42 is mutated to histidine, the enzyme develops a pH dependence corresponding to a loss of catalytic power upon deprotonation of the histidine. Structural data indicate that the change is not due to changes in active site architecture, but due to the difference in charge at this key site. With loss of the positive charge on the K42H sidechain at high pH, the enzyme retains lyase activity at approximately 100-fold lowered catalytic efficiency, but loses detectable mutase activity. We propose that both substrate organization and electrostatic transition state stabilization contribute to catalysis. However, the dominant reaction path for catalysis is dependent on reaction conditions, which influence the electrostatic properties of the enzyme active site amino acid sidechains.
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This publication was made possible by funds from the American Lung Association of Kansas and from the Kansas Masonic Cancer Research Institute, by the Graduate Training Program in Dynamic Aspects of Chemical Biology NIH grant T32 GM08545 (J.O.) from the National Institute of General Medical Sciences, by NIH grant P20 RR016475 from the INBRE Program of the National Center for Research Resources, and by NIH grants R01 AI77725 and K02 AI093675 from the National Institute for Allergy and Infectious Disease.
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2011-08-23
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Olucha, J., Ouellette, A. N., Luo, Q., & Lamb, A. L. (2011). pH Dependence of Catalysis by Pseudomonas aeruginosa Isochorismate-Pyruvate Lyase: Implications for Transition State Stabilization and the Role of Lysine 42. Biochemistry, 50(33), 7198–7207. http://doi.org/10.1021/bi200599j