Stability and Folding of Dihydrofolate Reductase from the Hyperthermophilic Bacterium Thermotoga maritima
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- 作者：Thomas Dams and Rainer Jaenicke
- 刊名：Biochemistry
- 出版年：1999
- 出版时间：July 13, 1999
- 年：1999
- 卷：38
- 期：28
- 页码：9169 - 9178
- 全文大小：114K
- 年卷期：v.38,no.28(July 13, 1999)
- ISSN：1520-4995
文摘
Dihydrofolate reductase (DHFR) has been a well-established model system for protein folding.The enzyme DHFR from the hyperthermophilic bacterium Thermotoga maritima (TmDHFR) displaysdistinct adaptations toward high temperatures at the level of both structure and stability. The enzymerepresents an extremely stable dimer; no isolated structured monomers could be detected in equilibriumor during unfolding. The equilibrium unfolding strictly follows the two-state model for a dimer (N2 
2U), with a free energy of stabilization of 
G = -142 ± 10 kJ/mol at 15 
C. The two-state model isapplicable over the whole temperature range (5-70 C), yielding a G vs T profile with maximum stabilityat around 35 C. There is no flattening of the stability profile. Instead, the enhanced thermostability ischaracterized 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. 108 times) compared to DHFR from Escherichia coli (EcDHFR).In contrast to EcDHFR, no evidence for heterogeneity of the native state is detectable. Refolding proceedsvia at least two intermediates and a burst-phase of rather low amplitude. Reassociation of monomericintermediates is not rate-limiting on the folding pathway due to the high association constant of the dimer.
2U), with a free energy of stabilization of G = -142 ± 10 kJ/mol at 15 C. The two-state model isapplicable over the whole temperature range (5-70 C), yielding a G vs T profile with maximum stabilityat around 35 C. There is no flattening of the stability profile. Instead, the enhanced thermostability ischaracterized 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. 108 times) compared to DHFR from Escherichia coli (EcDHFR).In contrast to EcDHFR, no evidence for heterogeneity of the native state is detectable. Refolding proceedsvia at least two intermediates and a burst-phase of rather low amplitude. Reassociation of monomericintermediates is not rate-limiting on the folding pathway due to the high association constant of the dimer.