Importance of Loop L1 Dynamics for Substrate Capture and Catalysis in Pseudomonas aeruginosa d -Arginine Dehydrogenase
| Autor: | Michael G. Souffrant, Giovanni Gadda, Sheena Vasquez, Donald Hamelberg, Daniel Ouedraogo |
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| Rok vydání: | 2017 |
| Předmět: |
Models
Molecular 0301 basic medicine Protein Folding Protein Conformation Stereochemistry Dehydrogenase Flavin group Molecular Dynamics Simulation Arginine Ligands Biochemistry Substrate Specificity 03 medical and health sciences Protein structure Bacterial Proteins Leucine Catalytic Domain Databases Protein Protein Unfolding chemistry.chemical_classification 030102 biochemistry & molecular biology biology Substrate (chemistry) Active site Stereoisomerism Hydrogen-Ion Concentration Recombinant Proteins 030104 developmental biology Enzyme chemistry Biocatalysis Mutation Pseudomonas aeruginosa Mutagenesis Site-Directed biology.protein Protein folding Amino Acid Oxidoreductases Oxidation-Reduction Algorithms |
| Zdroj: | Biochemistry. 56:2477-2487 |
| ISSN: | 1520-4995 0006-2960 |
| DOI: | 10.1021/acs.biochem.7b00098 |
| Popis: | Mobile loops located at the active site entrance in enzymes often participate in conformational changes required to shield the reaction from bulk solvent, to control the access of the substrate to the active site, and to position residues for substrate binding and catalysis. In d-arginine dehydrogenase from Pseudomonas aeruginosa (PaDADH), previous crystallographic data suggested that residues 45-47 in the FAD-binding domain and residues 50-56 in the substrate-binding domain in loop L1 could adopt two distinct conformations. In this study, we have used molecular dynamics, kinetics, and fluorescence spectroscopy on the S45A and A46G enzyme variants of PaDADH to investigate the impact of mutations in loop L1 on the catalytic function of the enzyme. Molecular dynamics showed that the mutant enzymes have probabilities of being in open conformations that are higher than that of wild-type PaDADH of loop L1, yielding an increased level of solvent exposure of the active site. In agreement, the flavin fluorescence intensity was ∼2-fold higher in the S45A and A46G enzymes than in wild-type PaDADH, with a 9 nm bathochromic shift of the emission band. In the variant enzymes, the k |
| Databáze: | OpenAIRE |
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