(655.12) Insights into the catalytic mechanism of M. tuberculosis indole-3-glycerol phosphate synthase

Poster Board Number: A181

Nina Goodey (Montclair State University), Sarah Cho (Montclair State University), Bhatti Maryum (Montclair State University), Huma Booter (Montclair State University), Cintya Moran (Montclair State University), Natalie Jefferson (Montclair State University), Savannah Van Den Berg (Montclair State University), Patryjca Marin (Montclair State University), Ashley Peralta (Montclair State University), Katherine Margolin (Montclair State University), David Konas (Montclair State University)

The pathogen M. tuberculosis causes the lung disease tuberculosis (TB). Emerging TB drug resistance worldwide indicates the need for new inhibitors and targets. Recent research suggests that the tryptophan biosynthetic pathway in M. tuberculosis can be targeted to treat TB. Indole-3-glycerol phosphate synthase (IGPS) in M. tuberculosis (MtIGPS) is an α/β barrel enzyme that catalyzes the fourth step in the tryptophan biosynthetic pathway: the conversion of 1-(o-carboxyphenylamino)-1-deoxyribulose 5’-phosphate (CdRP) into indole-glycerol-phosphate (IGP) via cyclization, decarboxylation and dehydration steps. The pH profile of MtIGPS was observed to have a bell shaped curve with pKa1 and pKa2 values or 6.7 ± 0.14 and 8.7 ± 0.12, supporting the involvement of both a general acid and a general base in MtIGPS catalysis.  Mutations were introduced to three residues that had been previously proposed to serve as the catalytic base: E57, E168 and E219. The E57Q variant had a gt;200-fold reduced catalytic activity compared to the wildtype and the E168Q variant had a gt;187-fold increased KM. The E219 sidechain interacts with K119, which is proposed to be the catalytic acid. The E219D variant had a gt;233-fold decreased catalytic activity. Combined with docking predictions, these data were consistent with E57 as the catalytic base, E168 playing a role in substrate binding, and E219 in orienting the catalytic acid K119. These data together provide an increased understanding of the structure-function relationship in the MtIGPS active site and insights in the IGPS enzyme family. 

Support or Funding Information

This work was supported by the National Institute of General Medical Sciences of the National Institutes of Health under award number R15GM126467 and by the National Science Foundation under award number 2030879.

lt;pgt;This work was supported by the National Institute of General Medical Sciences of the National Institutes of Health under award number R15GM126467 and by the National Science Foundation under award numberamp;nbsp;2030879.lt;/pgt;