Simplifying complex sequence information: A PCP-consensus protein binds antibodies against all four Dengue serotypes

Abstract

Designing proteins that reflect the natural variability of a pathogen is essential for developing novel vaccines and drugs. Flaviviruses, including Dengue (DENV) and West Nile (WNV), evolve rapidly and can “escape” neutralizing monoclonal antibodies by mutation. Designing antigens that represent many distinct strains is important for DENV, where infection with a strain from one of the four serotypes may lead to severe hemorrhagic disease on subsequent infection with a strain from another serotype. Here, a DENV physicochemical property (PCP)-consensus sequence was derived from 671 unique sequences from the Flavitrack database. PCP-consensus proteins for domain 3 of the envelope protein (EdomIII) were expressed from synthetic genes in E. coli. The ability of the purified consensus proteins to bind polyclonal antibodies generated in response to infection with strains from each of the four DENV serotypes was determined. The initial consensus protein bound antibodies from DENV-1-3 in ELISA and Western blot assays. This sequence was altered in 3 steps to incorporate regions of maximum variability, identified as significant changes in the PCPs, characteristic of DENV-4 strains. The final protein was recognized by antibodies against all four serotypes. Two amino acids essential for efficient binding to all DENV antibodies are part of a discontinuous epitope previously defined for a neutralizing monoclonal antibody. The PCP-consensus method can significantly reduce the number of experiments required to define a multivalent antigen, which is particularly important when dealing with pathogens that must be tested at higher biosafety levels.

D-graph clusters flaviviruses and β-coronaviruses according to their hosts, disease type and human cell receptors

“…Interestingly, Zika virus (node 18) falls exactly between the mosquito borne encephalitic and hemorrhagic groups, consistent with its cross reactivity with DENV antibodies (14). An identity matrix for the E protein domain 3 region of Zika compared to WNV, DENV strains and our 7p8 PCP-consensus protein that binds antibodies to all four DENV serotypes (15) illustrates how this Zika indeed lies between the encephalitic and hemorrhagic FV, with >50% identity to all ( Supplementary Fig. 1).…”

Sequence specificity for uridylylation of the viral peptide linked to the genome (VPg) of enteroviruses

“…It is designed to be useful for sets with many sequences (such as viral isolates) that have a high superficial redundancy (i.e., nearly identical sequences that differ at only a few positions). The rationale and details of the method are described in detail elsewhere (Bowen et al, 2012; Danecek et al, 2010; Danecek and Schein, 2010; Schein et al, 2012). Here, the PCP-con program was used to determine a consensus of 31 of the most diverse EV VPg sequences (Figure 1).…”

Cross-reactivities between human IgMs and the four serotypes of dengue virus as probed with artificial homodimers of domain-III from the envelope proteins

“…Table 1 gives the origin of the viral ED3 domain and the corresponding segment of the envelope protein. Table 2 gives the number of residue changes between the ED3 domains of any two DENV serotypes, and also between the ED3 domain of any DENV serotype and the consensus ED3 domain (DENVc) [63,64]. The productions of the H6-ED3-PhoA hybrids in the periplasmic space of E. coli and their purification from periplasmic extracts through their His-tag were performed essentially as described [41].…”