ABSTRACTMOLECULAR BASIS OF COMPLEMENT FACTOR-H RECRUITMENT BY THE LYME DISEASE PATHOGEN BORRELIA BURGDORFERIByJagannath SilwalLyme disease is the most common vector borne illness around the globe, caused by species of spirochetes in the Borrelia genus. In the United States, Borrelia burgdorferi sensu stricto (hereafter termed B. burgdorferi) is the major causative agent of Lyme disease, whereas Borrelia afzelii, Borrelia garinii and many other related Borrelia species are major pathogens responsible for Lyme disease in other parts of the world. B. burgdorferi is transmitted by hard-bodied tick species mainly from genus Ixodes and, like many other infectious pathogens, it has evolved many mechanisms to circumvent a highly sophisticated and tightly regulated host immune system, causing persistent infection. One of the key mechanisms that allows Borrelia to cause such infection is its ability to manipulate the host’s complement system by recruiting host regulator proteins to its own cell surface. The complement system is an integral part of the innate immune system, tightly regulated via interactions of several regulator proteins. The complement factor H (FH) protein is one of the key complement regulator proteins and plays a major role in avoiding complement attack on self-cells. FH consists of 20 complement control protein (CCP) domains and inhibits both complement activation and amplification on host cells by promoting inhibition and degradation of enzymes associated with innate immunity. B. burgdorferi recruits FH protein to its surface by expressing multiple surface proteins. This mechanism allows B. burgdorferi to evade host immune attack, leading to pathogen infections in the host. As the key immune evasion tool, B. burgdorferi expresses multiple FH-binding proteins on its surface during various stages of infection. So far, five different FH-binding surface proteins (CspA, CspZ, ErpA, ErpP and ErpC) are known in Borrelia. The structures of three of these surface proteins, CspA, CspZ and ErpA, in complex with different CCPs of human complement FH (hFH) were recently solved in our lab and are presented here.My research work focused on characterization of each of these three host-pathogen protein complexes using a combination of biochemical and biophysical methods. Interestingly, our results for CspA and CspZ protein complexes with hFH protein revealed unique binding sites in borrelial as well as FH protein, contradicting previous reports. Many results have been published in the last decades regarding the binding of FH with CspA protein, assigning 1) CspA dimer as the major FH binding site and 2) hFH CCP 7 as the predominant binding site for CspA. In contrast, our results show that hFH CCP 5 alone is sufficient for binding CspA, with nanomolar dissociation constant (Kd) in a different region from the previously published dimeric cleft in CspA. Similarly, hFH CCP 7 and CCP 20 bind to borrelial protein CspZ and ErpA, respectively. I will present the structures of all these protein complexes along with results from characterization of each of these protein complexes as determined by alanine scanning mutagenesis, isothermal titration calorimetry (ITC) and extensive computational analysis.
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