Chimeric Protein Engineering
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Protein stability can be enhanced by the incorporation of non-natural amino acids and semi-rigid peptidomimetics to lower the entropic penalty upon protein folding through preorganization. An example is the incorporation of aminoisobutyric acid (Aib, α-methylalanine) into proteins to restrict the Φ and Ψ backbone angles adjacent to Aib to those associated with helix formation. Reverse-turn analogs were introduced into the sequences of HIV protease and ribonuclease A that enhanced their stability and retained their native enzymatic activity. In this work, a chimeric protein, design_4, was engineered, in silico, by replacing the C-terminal helix of full sequence design protein (FSD-1) with a semi-rigid helix mimetic. Residues 1–16 of FSD-1 was ligated in silico with the N-terminus of a phenylbipyridyl-based helix mimetic to form design_4. The designed chimeric protein was stable and maintained the designed fold in a 100-nanosecond molecular dynamics simulation at 280 K. Its β-hairpin adopted conformations that formed three additional hydrogen bonds. Compared to FSD-1, design_4 contained fewer peptide bonds and internal degrees of freedom; it should, therefore, be more resistant to proteolytic degradation and denaturation.
Keywordsprotein engineering helix mimetic peptidomimetic chimeric protein
The authors would like to thank Daniel J. Kuster and Christy Taylor, Ph.D. for valuable discussions and critical reading of this manuscript. This research was supported in part by NIH research grant (GM 08460) to GRM. J.F. also acknowledges graduate support from the Division of Biology and Biomedical Science of Washington University in St. Louis, the Computational Biology Training Grant (GM 008802), and the Kauffman Foundation. Computational resources were supported in part by TeraGrid.
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