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Total Protein Extraction and 2-D Gel Electrophoresis Methods for Burkholderia Species
Published on: October 15, 2013
Changes in protein expression in Burkholderia vietnamiensis PR1 301 at pH 5 and 7 with and without nickel
Joy D Van Nostrand1, John M Arthur2, Lisa E Kilpatrick3
1Marine Biomedicine and Environmental Sciences Center, Medical University of South Carolina, 221 Fort Johnson Rd, Charleston, SC 29412, USA.
Abstract:
Burkholderia vietnamiensis PR1(301) (PR1) exhibits pH-dependent nickel (Ni) tolerance, with lower Ni toxicity observed at pH 5 than at pH 7. The Ni tolerance mechanism in PR1 is currently unknown, and traditional mechanisms of Ni resistance do not appear to be present. Therefore, 2D gel electrophoresis was used to examine changes in protein expression in PR1 with and without Ni (3.4 mM) at pH 5 and 7. Proteins with both a statistically significant and at least a twofold difference in expression level between conditions (pH, Ni) were selected and identified using MALDI-TOF-MS or LC-MS. Results showed increased expression of proteins involved in cell shape and membrane composition at pH 5 compared with pH 7. Scanning electron microscopy indicated elongated cells at pH 5 and 6 compared with pH 7 in the absence of Ni. Fatty acid methyl ester analysis showed a statistically significant difference in the percentages of long- and short-chain fatty acids at pH 5 and 7. These findings suggest that changes in membrane structure and function may be involved in the ability of PR1 to grow at higher concentrations of Ni at pH 5 than at pH 7.
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