Michelle Snyder

Structural Requirements for NADase activity of bacterial Toll/interleukin-1 receptor (TIR) domain-containing proteins part of CUREnet:CURE Collection
Toll-like receptors (TLRs) initiate innate immune signaling pathways via interactions of their Toll/interleukin-1 receptor (TIR) domains with cytoplasmic TIR domain-containing signaling proteins. Various bacterial species also express TIR domain-containing proteins that appear to contribute to bacterial evasion of the innate immune system. Bacterial TIR domains also have been found to exhibit NADase activity. We have developed a course-based undergraduate research experience (CURE class) involving 16-20 undergraduate students per year to characterize the structural requirements for the NADase activity of the TIR from Acetinobacter baumanii (AbTIR). Students use the molecular visualization software UCSF Chimera to analyze the AbTIR crystal structure and identify amino acids near the predicted NAD binding site in AbTIR that they hypothesize will play a role in AbTIR NADase activity. Students then perform site-directed mutagenesis to create plasmids for expression of their His-tagged mutant AbTIR proteins. Recombinant mutant AbTIR proteins are expressed in E. coli, purified by nickel chromatography and tested for NADase activity. Results from the course are presented at the annual department research poster symposium, allowing students a forum in which to practice their scientific communication skills and engage with faculty and other research students from the department. Overall, the course aims to provide research experiences for a large and diverse group of students, engaging these students in critical research skills including hypothesis formation, experimental design, data collection, data analysis, and communication of scientific findings.