Expression and Purification of a Novel Calcium-Binding Protein, EfhX, Necessary for Phytopathogenesis in Xanthomonas Strains
Poster Number
045
College
College of Arts and Sciences
Department
Chemistry, Physics, Geology, & the Environment
Faculty Mentor
Jason Hurlbert, Ph.D.
Abstract
We have identified a gene in several strains of Xanthomonas that, when expressed, can elicit a hypersensitive response (hr) in tomato plants. Hr elicitation is normally limited to specific bacterial-host pairings, but introduction and expression of this gene by bacterial species that do not normally infect tomato serves to elicit hr, indicating that the encoded protein is crucial to the infectious process. Bioinformatic analysis of the protein, which we have named EfhX (EF-Hand containing protein from Xanthomonas) reveals that the protein is predicted to contain a single transmembrane alpha-helix, spanning amino acids 60 to 81, and two calcium binding domains, termed EF-Hands, in the carboxy-terminal domain of the protein. In order to better understand the function of this novel protein, we have cloned the EfhX gene from Xanthomonas aurantofolia and expressed it in Escherichia coli so that we can obtain quantities of the protein sufficient to grow protein crystals and determine the structure of the protein via X-ray diffraction. We have successfully purified the protein to homogeneity as determined by SDS-PAGE and anti-hexahistidine Western Blot.
Previously Presented/Performed?
SAEOPP McNair/SSS Scholars Research Conference, Atlanta, Georgia, June 2018
Grant Support?
Supported by an SC INBRE grant from the National Institute for General Medical Sciences (NIH-NIGMS), and by a Ronald E. McNair Post-Baccalaureate Achievement Program grant from the U.S. Department of Education
Start Date
12-4-2019 2:15 PM
End Date
April 2019
Expression and Purification of a Novel Calcium-Binding Protein, EfhX, Necessary for Phytopathogenesis in Xanthomonas Strains
Richardson Ballroom – DiGiorgio Campus Center
We have identified a gene in several strains of Xanthomonas that, when expressed, can elicit a hypersensitive response (hr) in tomato plants. Hr elicitation is normally limited to specific bacterial-host pairings, but introduction and expression of this gene by bacterial species that do not normally infect tomato serves to elicit hr, indicating that the encoded protein is crucial to the infectious process. Bioinformatic analysis of the protein, which we have named EfhX (EF-Hand containing protein from Xanthomonas) reveals that the protein is predicted to contain a single transmembrane alpha-helix, spanning amino acids 60 to 81, and two calcium binding domains, termed EF-Hands, in the carboxy-terminal domain of the protein. In order to better understand the function of this novel protein, we have cloned the EfhX gene from Xanthomonas aurantofolia and expressed it in Escherichia coli so that we can obtain quantities of the protein sufficient to grow protein crystals and determine the structure of the protein via X-ray diffraction. We have successfully purified the protein to homogeneity as determined by SDS-PAGE and anti-hexahistidine Western Blot.
