An Ectromelia Virus Protein That Interacts with Chemokines through Their Glycosaminoglycan Binding Domain

Abstract

Poxviruses encode a number of secreted virulence factors that modulate the host immune response. The vaccinia virus A41 protein is an immunomodulatory protein with amino acid sequence similarity to the 35-kDa chemokine binding protein, but the host immune molecules targeted by A41 have not been identified. We report here that the vaccinia virus A41 ortholog encoded by ectromelia virus, a poxvirus pathogen of mice, named E163 in the ectromelia virus Naval strain, is a secreted 31-kDa glycoprotein that selectively binds a limited number of CC and CXC chemokines with high affinity. A detailed characterization of the interaction of ectromelia virus E163 with mutant forms of the chemokines CXCL10 and CXCL12␣ indicated that E163 binds to the glycosaminoglycan binding site of the chemokines. This suggests that E163 inhibits the interaction of chemokines with glycosaminoglycans and provides a mechanism by which E163 prevents chemokine-induced leukocyte migration to the sites of infection. In addition to interacting with chemokines, E163 can interact with high affinity with glycosaminoglycan molecules, enabling E163 to attach to cell surfaces and to remain in the vicinity of the sites of viral infection. These findings identify E163 as a new chemokine binding protein in poxviruses and provide a molecular mechanism for the immunomodulatory activity previously reported for the vaccinia virus A41 ortholog. The results reported here also suggest that the cell surface and extracellular matrix are important targeting sites for secreted poxvirus immune modulators.

Glycoprotein G from pseudorabies virus binds to chemokines with high affinity and inhibits their function

“…Mutation of a cluster of basic residues results in an hCXCL12a variant (mutant 3/6) unable to bind to GAGs. These mutants have been previously used to determine the chemokine domain involved in the interaction with another vCKBP, ectromelia virus E163 protein (Ruiz-Arguello et al, 2008). The number of response units (RU) obtained with the wt and the P2G mutant was similar (Fig.…”

“…Other vCKBPs also bind to both the receptor-and GAG-binding domains of the chemokines and inhibit chemotaxis in vitro (Bryant et al, 2003;Webb et al, 2003Webb et al, , 2004. However, inhibition of migration in vitro could not be shown for vCKBP that interact solely through the GAG-binding domain of the chemokine (Ruiz-Arguello et al, 2008;Seet & McFadden, 2002). These vCKBPs may impair chemotaxis in vivo but not in vitro through the inhibition of chemokine presentation by GAGs (Seet & McFadden, 2002).…”

Identification of the Pharmacophore of the CC Chemokine-binding Proteins Evasin-1 and -4 Using Phage Display

“…Binding the N-terminal region of chemokines appears to be a common feature of several CKBPs. For example, vCCI from several viruses, as well as M3 and CrmD bind the 20s loop of their respective ligands (11,12,16,19,20). In particular, the vCCIs bind to the conserved Arg or Lys equivalent to Arg-17 in CCL3.…”

“…CCL5 F12A was not used in the chemotaxis assay as the mutation of Phe-12, a pivotal residue involved in receptor activation (35), abrogates the chemotactic activity of the chemokine. Binding to both the mutants was significantly reduced: CCL5 has an affinity of 12 …”

“…Several studies aimed at identifying amino acids that are involved in the interaction between antagonistic binding proteins and their targets have revealed a number of critical residues on the surface of chemokines (11)(12)(13)(14)(15). The crystal struc-* This work was supported by the European Union Seventh Framework Programme (FP7-2007-2013) Grant HEALTH-F4 -2011-281608 TIMER (to P. B. and A. E. I. P.) and NIAID, National Institutes of Health Grant RO1AI37113 (to T. M. H.).…”