Evaluation of sequence variability in HIV-1 gp41 C-peptide helix-grafted proteins |
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Authors: | Rachel L Tennyson Susanne N Walker Terumasa Ikeda Reuben S Harris Brian R McNaughton |
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Institution: | 1. Department of Chemistry, Colorado State University, Fort Collins, CO, USA;2. Department of Biochemistry & Molecular Biology, Colorado State University, Fort Collins, CO, USA;3. Department of Biochemistry, Molecular Biology, and Biophysics, University of Minnesota, Minneapolis, MN, USA;4. Howard Hughes Medical Institute, University of Minnesota, Minneapolis, MN, USA |
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Abstract: | Many therapeutically-relevant protein-protein interactions (PPIs) have been reported that feature a helix and helix-binding cleft at the interface. Given this, different approaches to disrupting such PPIs have been developed. While short peptides (<15 amino acids) typically do not fold into a stable helix, researchers have reported chemical approaches to constraining helix structure. However, these approaches rely on laborious, and often expensive, chemical synthesis and purification. Our premise is that protein-based solutions that stabilize a therapeutically-relevant helix offer a number of advantages. In contrast to chemically constrained helical peptides, or minimal/miniature proteins, which must be synthesized (at great expense and labor), a protein can be expressed in a cellular system (like all current protein therapeutics). If selected properly, the protein scaffold can stabilize the therapeutically-relevant helix. We recently reported a protein engineering strategy, which we call “helix-grafted display”, and applied it to the challenge of suppressing HIV entry. We have reported helix-grafted display proteins that inhibit formation of an intramolecular PPI involving HIV gp41 C-peptide helix, and HIV gp41 N-peptide trimer, which contain C-peptide helix-binding clefts. Here, we used yeast display to screen a library of grafted C-peptide helices for N-peptide trimer recognition. Using ‘hits’ from yeast display library screening, we evaluated the effect helix mutations have on structure, expression, stability, function (target recognition), and suppression of HIV entry. |
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Keywords: | gp41 Helix HIV Protein engineering Protein evolution |
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