Elimination of a Misfolded Folding Intermediate by a Single Point Mutation
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- 作者：Jesper E. Mogensen ; Henrik Ipsen ; Jens Holm ; and Daniel E. Otzen
- 刊名：Biochemistry
- 出版年：2004
- 出版时间：March 30, 2004
- 年：2004
- 卷：43
- 期：12
- 页码：3357 - 3367
- 全文大小：265K
- 年卷期：v.43,no.12(March 30, 2004)
- ISSN：1520-4995
文摘
We present an analysis of the folding behavior of the 159-residue major birch pollen allergenBet v 1. The protein contains a water-filled channel running through it. Consequently, the protein has ahydrophobic shell, rather than a hydrophobic core. During the folding of the protein from either the urea-,pH-, or SDS-denatured state, Bet v 1 transiently populates a partially folded intermediate state. This stateappears to be misfolded, since it has to unfold at least partially to fold to the native state. The misfoldedintermediate is not, however, a result of the water-filled channel in Bet v 1. The intermediate completelydisappears in the mutant Tyr 
Trp120, in which the channel is still present. Tyr120 appears to behaveas a "negative gatekeeper" which attenuates efficient folding. The close structural homologue, the appleallergen Mal d 1, also folds without any detectable folding intermediates. However, the position of thetransition state on the reaction coordinate, which is a measure of its overall compactness relative to thedenatured and native states, is reduced dramatically from ca. 0.9 in Bet v 1 to around 0.5 in Mal d 1. Wesuggest that this large shift in the transition state structure is partly due to different local helix propensities.Given that individual mutations can have such large effects on folding, one should not a priori expectstructurally homologous proteins to fold by the same mechanism.
Trp120, in which the channel is still present. Tyr120 appears to behaveas a "negative gatekeeper" which attenuates efficient folding. The close structural homologue, the appleallergen Mal d 1, also folds without any detectable folding intermediates. However, the position of thetransition state on the reaction coordinate, which is a measure of its overall compactness relative to thedenatured and native states, is reduced dramatically from ca. 0.9 in Bet v 1 to around 0.5 in Mal d 1. Wesuggest that this large shift in the transition state structure is partly due to different local helix propensities.Given that individual mutations can have such large effects on folding, one should not a priori expectstructurally homologous proteins to fold by the same mechanism.