Abstract
Leptospirosis, which is caused by pathogen Leptospira, is the most common zoonotic disease emerged in the world. The organism enters the human body through mucous membranes or broken skin contact with the urine of an infected animal. Leptospira can cause damage of the kidney in the host and lead to tubulointerstitial nephritis. LipL41 is one of the major lipoprotein and important virulence factor located on the outer membrane of Leptosira. LipL41 was used to study the vaccines against human leptospirosis and the diagnostic detection, but the structural information is still unclear. Based on the sequence analysis of LipL41, we found out that the C-terminal and hydrophilic region of LipL41 (residue 256-355, denoted as LipL41-C100) contains five helices with two TPR motifs. Several studies show that TPR motif is involved in protein-protein interaction and oligomerlization. Analysis by circular dichroism (CD) showed that LipL41-C100 is a α-helix protein (48%). The signals in the 1H-15N HSQC spectrum of LipL41-C100 were broad and some of them were disappear and the observation represents that LipL41-C100 is a molten globule protein. Thermal shift assay, size-exclusion chromatography (SEC) and analytical ultracentrifugation (AUC) also revealed that LipL41-C100 has many common characteristics of molten globule. We further analyzed the folding porosity of LipL41 by Foldindex. The result predicted that LipL41 has an intrinsically unfolded domain (residue 256-355) which is the same as LipL41-C100. The flexible structure is related to the ligand binding ability. 2,2,2-Trifluoroethanol (TFE) have been shown to stabilize the helical structure of the protein. The CD spectrum revealed that the α-helix content of LipL41-C100 is increase from 48% to 75.9 % in the presence of 50% TFE. The NMR spectra of LipL41-C100 in the presence of 50% TFE also show that TFE stabilizes the regions in the molten globule state. The NMR experiments of LipL41-C100 are underway.