Fig. 1.

Construction of LCC‐expressing plasmid pHK‐LCC and extracellular expression of active LCC in C. thermocellum.

A. Construction of pHK‐LCC. SP, signal peptide, P₂₆₃₈, promoter sequence from gene Clo1313_2638, Ter, terminator, Rep Ori, the origin of replicon of E. coli, RepB, replicon origin of C. thermocellum, CAT, thiamphenicol resistant gene. The insertion of the construct ‘P₂₆₃₈‐SP‐lcc’ was verified by PCR with the primers seqF/R and the theoretical size of the PCR product was 1.3 kb. The gel analysis of the PCR product is shown on the right.

B. SDS‐PAGE analysis of cell lysates and extracellular proteins of DSM1313 and DSM1313:: pHK‐LCC stained with Coomassie Brilliant Blue.

C. Fast‐Red dye staining analysis of the protein gel against 1‐naphthyl acetate to visualize esterolytic activity. Lanes 1 and 2, culture supernatant of DSM1313 and DSM1313::pHK‐LCC respectively; lanes 3 and 4, cell lysate of DSM1313 and DSM1313::pHK‐LCC respectively. Ten µg protein was loaded in each lane.