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PMID:10413491
| Citation |
Dams, T and Jaenicke, R (1999) Stability and folding of dihydrofolate reductase from the hyperthermophilic bacterium Thermotoga maritima. Biochemistry 38:9169-78 |
|---|---|
| Abstract |
Dihydrofolate reductase (DHFR) has been a well-established model system for protein folding. The enzyme DHFR from the hyperthermophilic bacterium Thermotoga maritima (TmDHFR) displays distinct adaptations toward high temperatures at the level of both structure and stability. The enzyme represents an extremely stable dimer; no isolated structured monomers could be detected in equilibrium or during unfolding. The equilibrium unfolding strictly follows the two-state model for a dimer (N(2) right harpoon over left harpoon 2U), with a free energy of stabilization of DeltaG = -142 +/- 10 kJ/mol at 15 degrees C. The two-state model is applicable over the whole temperature range (5-70 degrees C), yielding a DeltaG vs T profile with maximum stability at around 35 degrees C. There is no flattening of the stability profile. Instead, the enhanced thermostability is characterized by shifts toward higher overall stability and higher temperature of maximum stability. TmDHFR unfolds in a highly cooperative manner via a nativelike transition state without intermediates. The unfolding reaction is much slower (ca. 10(8) times) compared to DHFR from Escherichia coli (EcDHFR). In contrast to EcDHFR, no evidence for heterogeneity of the native state is detectable. Refolding proceeds via at least two intermediates and a burst-phase of rather low amplitude. Reassociation of monomeric intermediates is not rate-limiting on the folding pathway due to the high association constant of the dimer. |
| Links |
PubMed Online version:10.1021/bi990635e |
| Keywords |
Bacterial Proteins/chemistry; Bacterial Proteins/metabolism; Enzyme Stability; Hot Temperature; Kinetics; Models, Chemical; Protein Folding; Spectrophotometry; Temperature; Tetrahydrofolate Dehydrogenase/chemistry; Tetrahydrofolate Dehydrogenase/metabolism; Thermotoga maritima/enzymology |
Significance
Annotations
| Gene product | Qualifier | GO Term | Evidence Code | with/from | Aspect | Extension | Notes | Status |
|---|---|---|---|---|---|---|---|---|
| GO:0004146: dihydrofolate reductase activity |
ECO:0000314: |
F |
The enzymes were not found to exist as monomers in equilibrium or while unfolding. This suggests that the protein is extremely stable throughout experimental temperatures from 5-70 degrees Celsius with a maximum stability at about 35 degrees C. |
complete | ||||
Notes
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References
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