Characterization of a Novel Cellulose Biosynthesis Inhibitor, CBI28, in Gluconacetobacter xylinus

To study the underlying mechanisms for microbial cellulose biosynthesis, a novel compound, CBI28, was used as an inhibitor along with classical genetics and EMS mutagenesis. An EZ-Link Biotin Hydrazide Kit was used to create a CBI28-Biotin conjugate for further studies. Gluconacetobacter xylinus cells were exposed to 10 muM CBI28 to induce cellulose biosynthesis inhibition, lysed and small hydrophobic molecules were extracted using methanol and Waters Oasis HLB SPE-Paks. Samples were separated and detected using the Ultra Performance Liquid Chromatograph-Mass Spectrometer/Photo Diode Array. Putative mutants were isolated but did not survive for further study. An ion with the expected mass of a CBI28-Biotin conjugate (552 m/z) was detected but not in sufficiently high concentrations for characterization. Metabolite studies revealed putative metabolites derived from the HLB SPE and methanol extractions with no significant difference in extraction methods. Potential metabolites with masses of ∼281.77 m/z and ∼79 m/z were detected in CBI28 exposed cells. Further analysis needs to be performed to determine if CBI28 metabolites prevent cellulose production.