Structure and Calcium-Binding Activity of LipL32, the Major Surface Antigen of Pathogenic Leptospira sp.

Leptospirosis, a spirochaetal zoonotic disease caused by Leptospira, has been recognized as an important emerging infectious disease. LipL32 is the major exposed outer membrane protein found exclusively in pathogenic leptospires, where it accounts for up to 75% of the total outer membrane proteins. It is highly immunogenic, and recent studies have implicated LipL32 as an extracellular matrix binding protein, interacting with collagens, fibronectin, and laminin. In order to better understand the biological role and the structural requirements for the function of this important lipoprotein, we have determined the 2.25-Å-resolution structure of recombinant LipL32 protein corresponding to residues 21–272 of the wild-type protein (LipL3221–272). The LipL3221–272 monomer is made of a jelly-roll fold core from which several peripheral secondary structures protrude. LipL3221–272 is structurally similar to several other jelly-roll proteins, some of which bind calcium ions and extracellular matrix proteins. Indeed, spectroscopic data (circular dichroism, intrinsic tryptophan fluorescence, and extrinsic 1-amino-2-naphthol-4-sulfonic acid fluorescence) confirmed the calcium-binding properties of LipL3221–272. Ca2+ binding resulted in a significant increase in the thermal stability of the protein, and binding was specific for Ca2+ as no structural or stability perturbations were observed for Mg2+, Zn2+, or Cu2+. Careful examination of the crystallographic structure suggests the locations of putative regions that could mediate Ca2+ binding as well as binding to other interacting host proteins, such as collagens, fibronectin, and laminin.