The effect of ion pairs on the thermal stability of D-glyceraldehyde 3-phosphate dehydrogenase from the hyperthermophilic bacterium Thermotoga maritima
| Autor: | Rainer Jaenicke, Andrea Tomschy, Gerald Böhm |
|---|---|
| Rok vydání: | 1994 |
| Předmět: |
Guanidinium chloride
Hot Temperature Stereochemistry Molecular Sequence Data Glutamic Acid Bioengineering Dehydrogenase Arginine Biochemistry Dissociation (chemistry) chemistry.chemical_compound Enzyme Stability Coenzyme binding Amino Acid Sequence Molecular Biology Glyceraldehyde 3-phosphate dehydrogenase chemistry.chemical_classification Aspartic Acid Gram-Negative Anaerobic Bacteria Base Sequence biology Chemistry Lysine Thermophile Glyceraldehyde-3-Phosphate Dehydrogenases NAD biology.organism_classification Enzyme Thermotoga maritima Mutagenesis Site-Directed biology.protein Sequence Alignment Biotechnology |
| Zdroj: | "Protein Engineering, Design and Selection". 7:1471-1478 |
| ISSN: | 1741-0134 1741-0126 |
| DOI: | 10.1093/protein/7.12.1471 |
| Popis: | D-Glyceraldehyde 3-phosphate dehydrogenase from Thermotoga maritima (TmGAPDH) is intrinsically thermostable, exhibiting a thermal transition beyond 105 degrees C. Neither the amino-acid composition nor homology modelling, based on sequence alignment and known 3-D structures of the enzyme from meso- and thermophiles, provide an explanation of the anomalous stability. Recent X-ray data suggest that an increased number of ion pairs is involved. To prove this hypothesis, a number of charged residues contributing to ion pairs in TmGAPDH, but absent in the moderately thermophilic enzyme were altered. Elimination of peripheral ion pairs (E103-K104, E261-R266) was found to be ineffective. Altering a central charge cluster (R10-D47, E314, D*186) led to a drastic decrease in coenzyme binding. As a consequence, guanidine-dependent deactivation is shifted to significantly lower guanidinium chloride (GdmCl) concentrations without altering the denaturation/dissociation profile of the wild type enzyme. Mutants in the S loop (R195D, R195D-D181K) lead to a biphasic profile in the GdmCl-dependent denaturation transition and significant destabilization; at room temperature no subunit dissociation could be observed. |
| Databáze: | OpenAIRE |
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