Circular permutation prediction reveals a viable backbone disconnection for split proteins: An approach in identifying a new functional split intein

Yun Tzai Lee, Tz Hsiang Su, Wei-Cheng Lo, Ping Chiang Lyu*, Shih Che Sue

*Corresponding author for this work

研究成果: Article同行評審

11 引文 斯高帕斯(Scopus)

摘要

Split-protein systems have emerged as a powerful tool for detecting biomolecular interactions and reporting biological reactions. However, reliable methods for identifying viable split sites are still unavailable. In this study, we demonstrated the feasibility that valid circular permutation (CP) sites in proteins have the potential to act as split sites and that CP prediction can be used to search for internal permissive sites for creating new split proteins. Using a protein ligase, intein, as a model, CP predictor facilitated the creation of circular permutants in which backbone opening imposes the least detrimental effects on intein folding. We screened a series of predicted intein CPs and identified stable and native-fold CPs. When the valid CP sites were introduced as split sites, there was a reduction in folding enthalpy caused by the new backbone opening; however, the coincident loss in entropy was sufficient to be compensated, yielding a favorable free energy for self-association. Since split intein is exploited in protein semi-synthesis, we tested the related protein trans-splicing (PTS) activities of the corresponding split inteins. Notably, a novel functional split intein composed of the N-terminal 36 residues combined with the remaining C-terminal fragment was identified. Its PTS activity was shown to be better than current reported two-piece intein with a short N-terminal segment. Thus, the incorporation of in silico CP prediction facilitated the design of split intein as well as circular permutants.

原文English
文章編號e43820
期刊PLoS ONE
7
發行號8
DOIs
出版狀態Published - 24 八月 2012

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