SELECTIVITY DETERMINANTS OF PEPTIDE-BINDING DOMAINS
BACTERIAL SORTASE ENZYMES
Bacterial sortases are cysteine transpeptidase enzymes that are responsible for covalently attaching proteins to the cell surface, e.g., to assemble pili. Sortases are also used in protein engineering, as they can covalently modify desired proteins with a wide variety of applications. Target recognition of the Class A sortase from Staphylococcus aureus is very specific, requiring an LPXTG sequence motif for initial cleavage. However, Class A sortases from other genera, e.g., Streptococcus, are more promiscuous. We are interested in using wild-type and chimeric proteins to understand sortase selectivity and target recognition through natural sequence variation, X-ray crystallography, and protein biochemistry.
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This work is supported by the NSF, CAREER grant CHE-2044958, NIH , grant 1R15GM154315-01 (Co-PIs Amacher, John Antos, and Jay McCarty), and the Research Corporation for Science Advancement Cottrell Scholars Award.
pdz DOMAINS
PDZ domains are important scaffolding domains in signaling and trafficking systems that bind the extreme C-terminus of target proteins. Although the PDZ interaction motif specifies critical residues for binding at two residues in target sequences, work by ourselves and others previously revealed that up to 6 residues are important for binding. We use protein biochemistry and structural biology to determine how specificity and selectivity is encoded in PDZ domains. We are also interested in whether or not these non-motif selectivity determinants are conserved in distantly related domains, specifically those from the single celled eukaryote, choanoflagellates. We have created two computational programs, MotifAnalyzer-PDZ and Domain Analysis and Motif Matcher (DAMM), which use sequence and structural conservation in multiple organisms in order to predict PDZ targets.
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This project was previously funded by the NSF, grant #1904711.
SH2 and SH3 domains
SH2 domains, which bind to phosphorylated tyrosine-containing sequences, and SH3 domains, which bind to poly-proline-containing sequences, are commonly found in tyrosine kinase signaling pathways. Target recognition in both domain families is encoded by structurally-conserved loops near the peptide-binding site. We are using chimeric proteins to investigate how these loops regulate selectivity.
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This project is funded by the NSF, CAREER grant #2044958.
