Adhesion of pathogenic Leplospira spp. to mammalian cells is mediated by their adhesins interacting with host cell receptors. In a previous study, we have identified two potential fibronectin (Fn) binding sites in central variable region (LigBCen) and C-terminal variable region (LigBCtv) of LigB, an adhesin of pathogenic Leptospira spp. In this study, we have further localized the Fn-binding site on LigBCen and found a domain of LigB (LigBCen2) (amino acids 10 14-1165) strongly bound to Fn. LigBCen2 bound to a 70 kDa domain of Fn including N-terminal domain (NTD) and gelatin binding domain (GBD), but with a higher binding affinity to NTD (K-d = 272 nM) than to GBD (Kd = 1200 nM). Except Fn, LigBCen2 also bound laminin and fibrinogen. LigBCen2 could bind MDCK cells, and blocked the binding of Leptospira on MDCK cells by 45%. These results suggest that LigBCon2 contributed to high affinity binding on NTD or GBD of Fn, laminin, and fibrinogen and mediated Leptospira binding on host cells. (c) 2007 Elsevier Inc. All rights reserved.