Mechanisms of lipoprotein sorting and secretion in borrelia spirochetes

The National Institute of Allergy and Infectious Diseases awarded the University of Kansas Medical Center Research Institute $782,462 on September 1, 2026, under the Allergy and Infectious Diseases Research program (CFDA 93.855) to investigate mechanisms of lipoprotein sorting and secretion in Borrelia spirochetes, the causative agents of Lyme disease and relapsing fever.

The research examines how Borrelia burgdorferi deploys surface lipoproteins—which cover the bacterium's unique double-membrane envelope and serve as virulence factors critical to transmission and pathogenesis. The work will identify and mechanistically characterize the periplasmic and outer-membrane components that enable these lipoproteins to transit from the periplasm to the bacterial surface, building on prior findings that surface lipoprotein secretion determinants localize to N-terminal disordered regions, that translocation requires partial unfolding, and that mature lipoproteins anchor in the outer-membrane surface leaflet.

Performance takes place in Kansas City, Kansas, with a period of performance from September 1, 2026, through August 31, 2027. This is a Project Grant, a standard assistance mechanism under the program supporting innovative research addressing emerging infectious disease challenges.

Abstract

Bacterial protein secretion is a fundamental physiological process that generates the cell envelope and maintains its integrity throughout the bacterial life cycle. In bacterial pathogens, a variety of protein secretion systems have been shown to deploy important virulence factors to the bacterial surface, into the milieu, or even directly into eukaryotic cells or other bacteria. Borrelia spirochetes, the causative agents of tick-borne lyme disease and relapsing fever, have a unique double-membrane envelope with periplasmic flagella. The borrelia surface lacks lipopolysaccharide and is instead covered by abundant, immunodominant and serotype-defining surface lipoproteins that serve as linchpins for transmission and pathogenesis. A recent study has shown that two thirds of the about 130 lipoproteins expressed by the lyme disease bacterium borrelia burgdorferi localize to the surface. Therefore, b. Burgdorferi is a perfect model organism for investigations into the secretion of bacterial surface lipoproteins. Several seminal studies have demonstrated that (i) borrelia surface lipoprotein secretion determinants commonly localize to n-terminal disordered tether regions of the mature lipoproteins, (II) translocation through the outer membrane can initiate at a lipoprotein's c terminus and requires an at least partially unfolded conformation, and (III) borrelia surface lipoproteins are ultimately anchored in the surface leaflet of the outer membrane bilayer. These data support the hypothesis that the borrelia surface lipoprotein secretion pathway includes a periplasmic mechanism that prevents premature folding of surface lipoprotein and an outer membrane translocon complex that allows for the flipping of lipoproteins from the periplasm to the surface. This proposal will test the above hypothesis by identifying and mechanistically defining the components of the b. Burgdorferi surface lipoprotein secretion pathway. Aim 1 will build on recent crispri gene silencing data and employ cryo-EM, site-directed mutagenesis, and quantitative proteomics to determine the structure- function relationships of a b. Burgdorferi OM protein shown to facilitate translocation of lipoproteins from the periplasm to the surface. Aim 2 will use crispri, x-RAY crystallography, site-directed mutagenesis, and quantitative proteomics to characterize the periplasmic proteins delivering surface lipoproteins from the inner membrane to the OM. Aim 3 will leverage time-lapse single-cell microscopy to track lipoprotein sorting and secretion pathway proteins throughout the cell cycle. This will define the sequestration of the b. Burgdorferi lipoproteome by both pathways in space and time, thereby elucidating the mechanistic underpinnings of sequential changes in surface lipoproteins that enables disseminated and chronic infection. Together, these experiments will use novel approaches to further elucidate how emerging pathogens of global importance generate and maintain their interface with the host. This will ultimately yield better tools for diagnostics and improved strategies for prevention and treatment.