Structural and functional studies of novel viral complement inhibitors

Abstract: Viruses have developed several strategies to evade the complement system, a part of the immune system. The fourth open reading frame of the human virus Kaposi's sarcoma-associated herpesvirus (KSHV) encodes a type 1 transmembrane protein. This contains four complement control protein domains (CCP domains), and is thus structurally similar to human complement inhibitors. We named it KSHV complement control protein (KCP). Using recombinant KCP, we showed that it can inhibit the complement system by disrupting a key enzymatic complex, the C3 convertase. We predicted the 3D structure of KCP by homology modeling. Functionally important sites in KCP were mapped using mutants where either one to three amino acids, or entire CCP domains, were altered. We also found that a patch of positively charged amino acids in CCP 1-2 of KCP can bind to heparin and weakly to heparin like proteoglycans at the surface of cells, which seems to aid the virus in the attachment step prior to cell entry. In order to study the biological role of complement inhibition for this type of virus, we want to use the KSHV homologue rhesus rhadinovirus (RRV), since this virus is both technically easier to work with than KCP and also permits in vivo studies, by naturally infecting Rhesus Macaques. RRV complement control protein (RCP) is the KCP homologue in RRV. We have characterized its gene, confirmed that RCP is expressed and present on the surface of virus and performed experiments indicating that it is a functional complement inhibitor.