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A single amino acid substitution alters ClpS2 binding specificity |
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| Authors: | Christina Bergonzo Kunal Dharmadhikari Emily Samuels Makenzie Christensen Jennifer Tullman |
| Affiliation: | 1. Biomolecular Structure and Function Group, Institute for Bioscience and Biotechnology Research, National Institute of Standards and Technology and University of Maryland, Rockville, Maryland, USA;2. Biomolecular Structure and Function Group, Institute for Bioscience and Biotechnology Research, National Institute of Standards and Technology and University of Maryland, Rockville, Maryland, USA Fischell Department of Bioengineering, University of Maryland, College Park, Maryland, USA;3. Fischell Department of Bioengineering, University of Maryland, College Park, Maryland, USA Department of Biological Sciences, University of Maryland, Rockville, Maryland, USA;4. Fischell Department of Bioengineering, University of Maryland, College Park, Maryland, USA Department of Biochemistry, Thomas S. Wootton High School, Rockville, Maryland, USA |
| Abstract: | ClpS2 is a small protein under development as a probe for selectively recognizing N-terminal amino acids of N-degron peptide fragments. To understand the structural basis of ClpS2 specificity for an N-terminal amino acid, all atom molecular dynamics (MD) simulations were conducted using the sequence of a bench-stable mutant of ClpS2, called PROSS. We predicted that a single amino acid leucine to asparagine substitution would switch the specificity of PROSS ClpS2 to an N-terminal tyrosine over the preferred phenylalanine. Experimental validation of the mutant using a fluorescent yeast-display assay showed an increase in tyrosine binding over phenylalanine, in support of the proposed hypothesis. |
| Keywords: | molecular dynamics protein engineering rational design |