G14secL04Tpc3

Lyme Disease and OspB protein

Symptoms and Origin of Lyme disease

Lyme disease is caused by the bacterial spirochete Borrelia burgdorferi. It is an inflammatory disorder that usually begins with skin lesions known as erythema chronicum migrands (ECM)^[1]. Months later the skin lesion can be followed by cardiac or neurological symptoms, migratory polyarthritis, oligoarticular arthritis, and chronic arthritis in the knees^[1]. The disease was first recognized as a new form of inflammatory arthritis in Lyme, Connecticut in 1975. Since then, the disease has been reported from many other parts of the United States^[1].

Outer Surface Proteins

Colonization and survival of Borrelia burgdorferi within ticks and mammals is facilitated, in part, by lipoproteins. OspA, OspB, and OspC are three of the major lipoproteins present on the outer surface of the spirochete Borrelia burgdoferi. Studies have shown that OspA and OspB are critical for the adherence of the spirochete to the gut wall of its tick vector^[2]. The free OspB structure consists of a barrel domain which might be the portion that interacts with a protein or a linear saccharide in the tick-gut, promoting the attachment of spirochete on the tick gut^[3]. It’s speculated that destroying these lipoproteins will cause lysis of spirochetes. Some antibody Fab fragments such as H6831 and CB2 have been shown to cause bacterial lysis of spirochetes by binding to these lipoproteins in the absence of phagocytes and without complement, which are normally part of the immune response to bacteria^[3]. However, it remains unclear how binding of H6831 or CB2 can lead directly to lysis of the bacterium.

Transmission of Spirochete

Selective expression of outer surface proteins are important for the colonization and persistence within the tick vector. It was observed that after entry into the ticks, B. burgdorferi replicates and persists within the gut, then during a subsequent blood meal, migrates through the vector and is transmitted to a new host.^[4]. Other studies show that the expression of OspA and OspB is immediately turned on when the spirochetes enter and reside within the arthropod vector^[4]. However, during transmission from the arthropod vector to a vertebrate host, expression of OspA and OspB are downregulated while OspC is upregulated ^[4]. Since OspA and OspB are critical for the adherence of the spirochete to the mid-gut of the tick, if these proteins are down-regulated or expressed less, then the spirochete will attach less to the mid-gut of the tick and as a result be more likely to be excreted in some way into the host on which the tick is feeding.

Free OspB structure

structure consists of twelve anti-parallel beta-strands followed by a single alpha helix. and form two sheets that bend, along with the final that comes over the top into the . It has been shown that OspB deficient spirochetes lack the ability to bind to the tick gut ^[2]. Further studies suggest that the structural barrel domain is what gives OspB the ability to adhere to the tick gut wall ^[3]. The barrel domain cavity could serve as the binding site for an exposed protein loop, small peptide, or linear saccharide hanging off the tick gut wall^[3]. on free OspB's N-terminus are cleaved when OspB is bound to antibody fragment H6831, suggesting that they're removed by proteolysis in the H6831-bound complex^[3]. The OspB protein is an integral membrane lipoprotein, with its lipidated N-terminus imbedded in the outer membrane of the spirochete and its C-terminus directed away from the membrane^[5]. Therefore, it is speculated that proteolytic action on these N-terminus beta strands somehow cause lysis of the spirochete. Studies have shown that when complement-independent antibodies such as H6831 and CB2 act on OspB, the result is blebbing of the outer membrane of the bacteria ^[6]. This blebbing of the bacterial membrane has been shown to create small openings in the outer membrane of the bacteria, which leads for potential osmotic lysis depending on the solution the bacteria is in^[6].

OspB complexed with H6831 Antibody Fab Fragment

(PDB code 1rjl).

( and ) has been shown to bind on an epitope of the ^[3]. Once this occurs, the resulting OspB∙H6831 complex is formed and the first four beta strands of the OspB protein are proteolysed^[3]. Cleavage of these first four beta strands somehow causes bacterial lysis of the B. burdgorferi. This loss is the most significant conformational change in the OspB protein itself, and all other changes are minor and appear to be related to this loss. The majority of the binding occurs on the C-terminus of OspB. The C-terminus consists of

that are involved in the electrostatic forces keeping the H6831 antibody bound to the OspB protein^[3]. Of these three loops, plays the most significant role in keeping the complex together. This is due to the presence of a on this loop. This important lysine forms an ionic salt bridge with the on the heavy chain of the antibody Fab Fragment. Thus, formation of the complex consists of the C-terminus of the OspB protein coming into contact with the variable regions of the H6831 antibody Fab fragment and the lysine-253 wedging between the two (tryptophan-33 and tyrosine-101) and salt bridging with the glu-50 on the heavy chain of the antibody^[3]. Studies show that other bacterial strains having a cysteine, glycine, glutamic acid, or threonine in place of Lys-253 demonstrate resistance to the bactericidal effects of H6831 and also less binding affinity for the antibody Fab fragment ^[7]. This bolsters how important the Lys-253 is in the binding between the OspB protein and the antibody. Additionally, one report found that an ospB escape mutant in a population of infectious B. burgdorferi, with a single base change in the ribosomal binding sequence and a single nucleotide deletion in the open reading frame of the ospB gene, demonstrated reduced expression and truncation of the OspB protein^[8]. This reduced expression and truncation of OspB extenuated the penetration capability and infectivity of the spirochete in the human umbilical vein endothelium cells^[8]. Thus, the OspB protein plays a fairly important role in the ability of the spirochete to infect a host, given that when it is damaged the spirochete's ability to infect the host is mitigated. After the complex is formed and the first four beta strands are cleaved off, a relic of this loss is a single on the N-terminus of the protein. This beta strand plays a role in the formation of dimers of OspB∙H6831 complex ^[3] during crystallization. Although the middle loop bearing the lysine is very important, the other loops certainly contribute to the bonding between OspB and the antibody fab fragment. For example, a at residue 276 on another loop interacts with residues on the light chain of the antibody fragment.