Fig 1.
The affinity of chFVN145 for recombinant protein E of TBEV.
Serial three-fold dilutions of chFVN145 starting from 81 nM were used as the analyte. Fitted traces are depicted as black lines. A global analysis of the interaction demonstrated a good quality fit and the affinity constant, calculated as KD = kd/ka, was 1.5 ± 0.2 nM.
Fig 2.
Post-exposure protection of mice infected with various subtypes of TBEV.
BALB/c mice (10–11 g) were treated (i.v.) with chFVN145 or anti-TBE-Ig at the indicated doses one day after infection with (A) 159 LD50 TBEV-Eu, (B) 231 LD50 TBEV-FE, and (C) 251 LD50 TBEV-Sib.
Fig 3.
Therapeutic efficacy of chFVN145 in mice infected with different subtypes of TBEV.
BALB/c mice (10–11 g) were treated (i.v.) once with chFVN145 or anti-TBE-Ig one, two, and three days after infection with (A) 20 LD50 TBEV-Eu, (B) 501 LD50 TBEV-FE, and (C) 3981 LD50 TBEV-Sib.
Fig 4.
Determination of non-protective doses of chFVN145 doses.
BALB/c mice (10–11 g) were treated (i.v.) with chFVN145 at the indicated doses one day after infection with (A) 199.5 LD50 TBEV and (B) 316 LD50 TBEV.
Fig 5.
The mean survival time of mice treated with non-protective doses of chFVN145.
The box-and-whisker plots represent the mean survival time of different groups of mice infected with TBEV with 100% mortality rate. Administration of non-protective doses did not decrease the survival rate and MST of treated mice compared to those of mice received 0.9% NaCl and non-treated controls independent of the timing of chFVN145 injection and TBEV strain used for infection.
Fig 6.
(A) Western blot analysis of lysates of E. coli cells producing rED1+2 (1), rED3_301 (2), rED3delD+ (3) and rED3delA+ (4), which were fractionated by SDS-PAGE (12.5%) and developed with chFVN145. Protein marker masses, in kilodaltons, are shown at the left side of the gel. (B) Results of phage display screening for peptides bound by chFVN145. Alignment of TBEV glycoprotein E homolog fragments and peptides, selected from PhD-12 and PhDC7C library TBEV protein E-like aa was marked in grey. (C) Ribbon and surface representations of TBEV glycoprotein E dimer structure (PDB 1svb) with indicated putative epitope sites. View from the outer surface of virion. (D) Ribbon and surface representations of proposed epitope sites, view from the DIII side of E glycoprotein. Molecular coordinates for the TBEV glycoprotein E structure (PDB 1svb) used in structural analysis were obtained from the Protein Data Bank then visualized using PyMol software (version 1.7.6.0).