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Improving the anti-toxin abilities of the CMG2-Fc fusion protein with the aid of computational design.

Xi Y, Wu X, Gao L, Shao Y, Peng H, Chen H, Chen H, Hu X, Yue J - PLoS ONE (2014)

Bottom Line: An experimental affinity assay revealed that the two variants showed increased binding affinity, and in vitro and in vivo toxin neutralization testing indicated that one of these mutants (CMG2-Fc(E117Q)) has superior activity against anthrax toxin and was suitable for further development as a therapeutic agent for anthrax infections.This study shows that the computational design of the PA binding interface of CMG2 to obtain CMG2-Fc variants with improving anti-toxin abilities is viable.Our results demonstrate that computational design can be further applied to generate other CMG2-Fc mutants with greatly improved therapeutic efficacy.

View Article: PubMed Central - PubMed

Affiliation: Beijing Institute of Biotechnology, Beijing, China.

ABSTRACT
CMG2-Fc is a fusion protein composed of the extracellular domain of capillary morphogenesis protein 2 (CMG2) and the Fc region of human immunoglobulin G; CMG2-Fc neutralizes anthrax toxin and offers protection against Bacillus anthracis challenge. To enhance the efficacy of CMG2-Fc against anthrax toxin, we attempted to engineer a CMG2-Fc with an improved affinity for PA. Using the automatic design algorithm FoldX and visual inspection, we devised two CMG2-Fc variants that introduce mutations in the CMG2 binding interface and improve the computationally assessed binding affinity for PA. An experimental affinity assay revealed that the two variants showed increased binding affinity, and in vitro and in vivo toxin neutralization testing indicated that one of these mutants (CMG2-Fc(E117Q)) has superior activity against anthrax toxin and was suitable for further development as a therapeutic agent for anthrax infections. This study shows that the computational design of the PA binding interface of CMG2 to obtain CMG2-Fc variants with improving anti-toxin abilities is viable. Our results demonstrate that computational design can be further applied to generate other CMG2-Fc mutants with greatly improved therapeutic efficacy.

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The effect of CMG2-Fc (E117Q) on the survival of F344 rats challenged with lethal toxin (LeTx).The weight ratio of CMG2-Fc to lethal factor was evaluated at 1∶1. The rest may be deduced by analogy.
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pone-0104674-g005: The effect of CMG2-Fc (E117Q) on the survival of F344 rats challenged with lethal toxin (LeTx).The weight ratio of CMG2-Fc to lethal factor was evaluated at 1∶1. The rest may be deduced by analogy.

Mentions: The ability of CMG2-Fc (E117Q) and CMG2-Fc to neutralize LeTx was compared in vivo. Male Fisher 344 rats were injected via the tail vein with LeTx (100 µg PA and 100 µg LF), either alone or in combination with various doses CMG2-Fc (E117Q) (100 µg, 50 µg or 25 µg). The results (Figure 5) show that all of the rats that received only LeTx died within 2 hours; however, for the CMG2-Fc (E117Q) groups, all of the rats receiving LeTx and CMG2-Fc (E117Q) at the mass ratio of 0.5∶1 survived, and only one rat died at the lower ratio of CMG2-Fc(E117Q) to LeTx (0.25∶1). In contrast, three of the rats receiving LeTx and CMG2-Fc at a ratio of 0.5∶1 died, and three of the rats receiving the lower ratio of CMG2-Fc:LeTx (0.25∶1) also died (Figure 5).


Improving the anti-toxin abilities of the CMG2-Fc fusion protein with the aid of computational design.

Xi Y, Wu X, Gao L, Shao Y, Peng H, Chen H, Chen H, Hu X, Yue J - PLoS ONE (2014)

The effect of CMG2-Fc (E117Q) on the survival of F344 rats challenged with lethal toxin (LeTx).The weight ratio of CMG2-Fc to lethal factor was evaluated at 1∶1. The rest may be deduced by analogy.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4125234&req=5

pone-0104674-g005: The effect of CMG2-Fc (E117Q) on the survival of F344 rats challenged with lethal toxin (LeTx).The weight ratio of CMG2-Fc to lethal factor was evaluated at 1∶1. The rest may be deduced by analogy.
Mentions: The ability of CMG2-Fc (E117Q) and CMG2-Fc to neutralize LeTx was compared in vivo. Male Fisher 344 rats were injected via the tail vein with LeTx (100 µg PA and 100 µg LF), either alone or in combination with various doses CMG2-Fc (E117Q) (100 µg, 50 µg or 25 µg). The results (Figure 5) show that all of the rats that received only LeTx died within 2 hours; however, for the CMG2-Fc (E117Q) groups, all of the rats receiving LeTx and CMG2-Fc (E117Q) at the mass ratio of 0.5∶1 survived, and only one rat died at the lower ratio of CMG2-Fc(E117Q) to LeTx (0.25∶1). In contrast, three of the rats receiving LeTx and CMG2-Fc at a ratio of 0.5∶1 died, and three of the rats receiving the lower ratio of CMG2-Fc:LeTx (0.25∶1) also died (Figure 5).

Bottom Line: An experimental affinity assay revealed that the two variants showed increased binding affinity, and in vitro and in vivo toxin neutralization testing indicated that one of these mutants (CMG2-Fc(E117Q)) has superior activity against anthrax toxin and was suitable for further development as a therapeutic agent for anthrax infections.This study shows that the computational design of the PA binding interface of CMG2 to obtain CMG2-Fc variants with improving anti-toxin abilities is viable.Our results demonstrate that computational design can be further applied to generate other CMG2-Fc mutants with greatly improved therapeutic efficacy.

View Article: PubMed Central - PubMed

Affiliation: Beijing Institute of Biotechnology, Beijing, China.

ABSTRACT
CMG2-Fc is a fusion protein composed of the extracellular domain of capillary morphogenesis protein 2 (CMG2) and the Fc region of human immunoglobulin G; CMG2-Fc neutralizes anthrax toxin and offers protection against Bacillus anthracis challenge. To enhance the efficacy of CMG2-Fc against anthrax toxin, we attempted to engineer a CMG2-Fc with an improved affinity for PA. Using the automatic design algorithm FoldX and visual inspection, we devised two CMG2-Fc variants that introduce mutations in the CMG2 binding interface and improve the computationally assessed binding affinity for PA. An experimental affinity assay revealed that the two variants showed increased binding affinity, and in vitro and in vivo toxin neutralization testing indicated that one of these mutants (CMG2-Fc(E117Q)) has superior activity against anthrax toxin and was suitable for further development as a therapeutic agent for anthrax infections. This study shows that the computational design of the PA binding interface of CMG2 to obtain CMG2-Fc variants with improving anti-toxin abilities is viable. Our results demonstrate that computational design can be further applied to generate other CMG2-Fc mutants with greatly improved therapeutic efficacy.

Show MeSH
Related in: MedlinePlus