Cell-surface protein YwfG ofLactococcus lactisbinds to α-1,2-linked mannose

Abstract

Lactococcus lactis strains are used as starter cultures in the production of fermented dairy and vegetable foods, but the species also occurs in other niches such as plant material. Lactococcus lactis subsp. lactis G50 (G50) is a plant-derived strain and potential candidate probiotics. Western blotting of cell-surface proteins using antibodies generated against whole G50 cells identified a 120-kDa protein. MALDI-TOF MS analysis identified it as YwfG, a Leu-Pro-any-Thr-Gly cell wall anchor domain–containing protein. Based on a predicted structure, a recombinant protein covering the N-terminal half (aa 28–511) of YwfG (YwfG28−511) was crystallized and the crystal structure was determined. The structure consisted of an L-type lectin domain, a mucin-binding protein domain, and a mucus-binding protein repeat. Recombinant proteins containing combinations of these domains were prepared and their interactions with monosaccharides were examined by isothermal titration calorimetry; the only interaction observed was between the L-type lectin domain and D-mannose. Among four mannobioses, α1,2-mannobiose had the highest affinity for the L-type lectin domain (dissociation constant = 34 μM). The L-type lectin domain also interacted with yeast mannoproteins and yeast cells. Soaking of the crystals of YwfG28−511 with mannose or α1,2-mannobiose revealed that both sugars bound to the L-type lectin domain in a similar manner, although the presence of the mucin-binding protein domain and the mucus-binding protein repeat within the recombinant protein inhibited the interaction between the L-type lectin domain and mannose. These findings provide new structural and functional insights into the interaction between L. lactis and its ecological niche via binding of the cell-surface protein YwfG with mannose.