Role of Inhibitor Aliphatic Chain in the Thermodynamics of Inhibitor Binding to Escherichia coli Enoyl-ACP Reductase and the Phe203Leu Mutant: A Proposed Mechanism for Drug Resistance
| Autor: | Christie G. Brouillette, Steve Dunham, Karen Siegel, Mark E. Snow, Irina I. Protasevich, J. Ronald Rubin, Rocco Gogliotti |
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| Rok vydání: | 2004 |
| Předmět: |
Protein Folding
Hot Temperature Protein Conformation Stereochemistry Entropy Phenylalanine Mutant Calorimetry Reductase medicine.disease_cause Biochemistry Structure-Activity Relationship Phenols Leucine Drug Resistance Bacterial medicine Point Mutation Enzyme Inhibitors Ternary complex Escherichia coli chemistry.chemical_classification Binding Sites Chemistry Escherichia coli Proteins Cooperative binding Isothermal titration calorimetry Enoyl-(Acyl-Carrier-Protein) Reductase (NADH) Recombinant Proteins Enzyme Thermodynamics Fatty Acid Synthases Oxidoreductases |
| Zdroj: | Biochemistry. 43:13380-13389 |
| ISSN: | 1520-4995 0006-2960 |
| DOI: | 10.1021/bi0492509 |
| Popis: | The antibacterial target enoyl-acyl carrier protein (ACP) reductase is a homotetrameric enzyme that catalyzes the last reductive step of fatty acid biosynthesis. In the present paper, four 2-(2-hydroxyphenoxy)phenol inhibitors, wherein the 4-position substituent varied from H to n-propyl, were studied to determine the contribution of the aliphatic chain to the binding to the wild-type (wt) enoyl-ACP reductase from Escherichia coli (FabI) and a drug-resistant mutant, (F203L)FabI, in which phenylalanine 203 is mutated to leucine. Thermodynamic parameters of ternary complex formation (enzyme-NAD(+)-inhibitor) were determined by isothermal titration calorimetry. The inhibitor affinity to wt FabI and (F203L)FabI increases with increasing aliphatic chain length, although the corresponding affinity for (F203L)FabI is lower, and also, it shows no detectable binding to the 4-H inhibitor. A distinguishing feature of inhibitor binding to either binary enzyme-NAD(+) complex is the apparent negative cooperativity for binding to the tetramer with half-site occupancy. For both enzymes, binding is enthalpy, DeltaH, driven. However, binding DeltaH becomes less favorable with increasing aliphatic chain length. Increases in affinity are found to be exclusively due to favorable changes in solvation entropy. Incremental changes in thermodynamic parameters within the series of inhibitors binding to wt FabI and (F203L)FabI are approximately the same. However, absolute differences between the two enzymes for binding to a given inhibitor are significant, suggesting different binding modes. This finding, coupled with a binding site conformation that is likely to be more rigid in the mutant, appears to result in the drug resistance of (F203L)FabI. |
| Databáze: | OpenAIRE |
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