OBJECTIVE: The aim of the present study was to compare the binding activity of Lactococcus lactis peptidoglycan protein anchor (PA) with different number of motifs. METHODS: L. lactis PA gene sequences with 2 and 3 lysin motifs (LysM) were obtained by PCR amplification. Then, the recombinant plasmid of pET-32a(+) containing PA gene was constructed and transformed into E. coli BL21 (DE3) , and induced to express the fusion protein PA2 and PA3. The purified and refolded PA2 and PA3 were incubated with gram positive enhancer matrix (GEM). The binding activities of PA2 and PA3 were identified and compared by Western blot, TEM and SDS-PAGE. RESULTS: The PA2 and PA3 could bind to GEM. The anchoring activity of PA3 was obviously superior to the PA2. CONCLUSION: The data indicated that PA with 3 LysMs had a better binding capacity compared with 2 LysMs. The results provided foundations to further improve the design of GEM-PA display system.
OBJECTIVE: The aim of the present study was to compare the binding activity of Lactococcus lactis peptidoglycan protein anchor (PA) with different number of motifs. METHODS:L. lactisPA gene sequences with 2 and 3 lysin motifs (LysM) were obtained by PCR amplification. Then, the recombinant plasmid of pET-32a(+) containing PA gene was constructed and transformed into E. coli BL21 (DE3) , and induced to express the fusion protein PA2 and PA3. The purified and refolded PA2 and PA3 were incubated with gram positive enhancer matrix (GEM). The binding activities of PA2 and PA3 were identified and compared by Western blot, TEM and SDS-PAGE. RESULTS: The PA2 and PA3 could bind to GEM. The anchoring activity of PA3 was obviously superior to the PA2. CONCLUSION: The data indicated that PA with 3 LysMs had a better binding capacity compared with 2 LysMs. The results provided foundations to further improve the design of GEM-PA display system.