Chitin recognition by a periplasmic chitooligosaccharide-binding protein from Vibrio cholerae.

The mechanism of chitin recognition by a periplasmic chitooligosaccharide-binding protein from Vibrio cholerae (VcCBP) was studied by thermal shift assays and isothermal titration calorimetry using di-N-acetylchitobiose, (GlcNAc)2; mono-N-acetylchitobioses, GlcN-GlcNAc and GlcNAc-GlcN; and fully de-N-acetylated chitobiose, (GlcN)2; as the ligands. As judged from the thermal shifts (ΔTm) of VcCBP upon the addition of individual chitobioses, the binding abilities toward VcCBP appeared to decrease in the order of (GlcNAc)2 > GlcN-GlcNAc > GlcNAc-GlcN ≫ (GlcN)2. Although the de-N-acetylation effect of the reducing end GlcNAc was more significant than that of the non-reducing end, both N-acetyl groups were found to cooperatively contribute to the interaction between VcCBP and (GlcNAc)2. The binding affinity of GlcN-GlcNAc to VcCBP was lower than that of (GlcNAc)2 by only 0.5 kcal·mol-1 of ΔG°; however, the entropy gain (-TΔS°) was enhanced in the former compared with the latter. GlcN-GlcNAc are likely to bind loosely to VcCBP but unlikely to undergo translocation by the VcCBP-mediated transporter system.