NMR and Crystal Structures of the Pyrococcus horikoshii RadA Intein Guide a Strategy for Engineering a Highly Efficient and Promiscuous Intein
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چکیده
منابع مشابه
NMR and crystal structures of the Pyrococcus horikoshii RadA intein guide a strategy for engineering a highly efficient and promiscuous intein.
In protein splicing, an intervening protein sequence (intein) in the host protein excises itself out and ligates two split host protein sequences (exteins) to produce a mature host protein. Inteins require the involvement for the splicing of the first residue of the extein that follows the intein (which is Cys, Ser, or Thr). Other extein residues near the splicing junctions could modulate splic...
متن کاملCloning, expression, purification, crystallization and preliminary X-ray diffraction data of the Pyrococcus horikoshii RadA intein.
The RadA intein from the hyperthermophilic archaebacterium Pyrococcus horikoshii was cloned, expressed and purified for subsequent structure determination. The protein crystallized rapidly in several conditions. The best crystals, which diffracted to 1.75 Å resolution, were harvested from drops consisting of 0.1 M HEPES pH 7.5, 3.0 M NaCl and were cryoprotected with Paratone-N before flash-cool...
متن کاملA promiscuous split intein with expanded protein engineering applications.
The protein trans-splicing (PTS) activity of naturally split inteins has found widespread use in chemical biology and biotechnology. However, currently used naturally split inteins suffer from an "extein dependence," whereby residues surrounding the splice junction strongly affect splicing efficiency, limiting the general applicability of many PTS-based methods. To address this, we describe a m...
متن کاملEvaluation and comparison of protein splicing by exogenous inteins with foreign exteins in Escherichia coli.
Protein splicing catalyzed by inteins has enabled various biotechnological applications such as protein ligation. Successful applications of inteins are often limited by splicing efficiency. Here, we report the comparison of protein splicing between 20 different inteins from various organisms in identical contexts to identify robust inteins with foreign exteins. We found that RadA intein from P...
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ژورنال
عنوان ژورنال: Journal of Molecular Biology
سال: 2012
ISSN: 0022-2836
DOI: 10.1016/j.jmb.2012.04.029