Cellulosomic profiling produced by Clostridium cellulovorans during growth on different carbon sources explored by the cohesin marker

Clostridium cellulovorans produces large extracellular enzyme complex, called cellulosomes. The diversity of the cellulosomal enzymes, which are secreted by C. cellulovorans that has been cultured on different carbon sources, such as Avicel, xylan, AXP (Avicel-xylan-pectin, 3:1:1) and cellobiose, was explored by two-dimensional gel electrophoresis. To identify the cellulosomal enzymes, we constructed a biomarker using cohesin 6, one of the CbpA cohesins, that was labeled with fluorescence. The major apparent spots were isolated and identified by ESI MS/MS protein sequencing. Fluorescently labeled cohesin clearly showed that the amount of the cellulosomal enzymes was influenced by the available carbon source. EngE, ExgS, EngK, XynB and ManA were most frequently expressed under all conditions. However, EngY was only observed on the AXP culture. We found two novel putative cellulosomal proteins, NC1[GH9] and NC2[GH26], and five unknown proteins, NU1, NU2, NU3, NU4 and NU5. The cohesin biomarker clearly showed different production patterns of the cellulosomal subunits under different culture conditions and revealed novel cellulosomal subunits.