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A truncated receptor-binding domain of MERS-CoV spike protein potently inhibits MERS-CoV infection and induces strong neutralizing antibody responses: implication for developing therapeutics and vaccines.

Du L, Kou Z, Ma C, Tao X, Wang L, Zhao G, Chen Y, Yu F, Tseng CT, Zhou Y, Jiang S - PLoS ONE (2013)

Bottom Line: Development of effective therapeutics and vaccines is crucial to save lives and halt the spread of MERS-CoV.The recombinant S377-588-Fc is able to induce in the vaccinated mice strong MERS-CoV S-specific antibodies, which blocks the binding of RBD to DPP4 receptor and effectively neutralizes MERS-CoV infection.These findings indicate that this truncated RBD protein shows promise for further development as an effective and safe vaccine for the prevention of MERS-CoV infection.

View Article: PubMed Central - PubMed

Affiliation: Lindsley F. Kimball Research Institute, New York Blood Center, New York, New York, United States of America.

ABSTRACT
An emerging respiratory infectious disease with high mortality, Middle East respiratory syndrome (MERS), is caused by a novel coronavirus (MERS-CoV). It was first reported in 2012 in Saudi Arabia and has now spread to eight countries. Development of effective therapeutics and vaccines is crucial to save lives and halt the spread of MERS-CoV. Here, we show that a recombinant protein containing a 212-amino acid fragment (residues 377-588) in the truncated receptor-binding domain (RBD: residues 367-606) of MERS-CoV spike (S) protein fused with human IgG Fc fragment (S377-588-Fc) is highly expressed in the culture supernatant of transfected 293T cells. The purified S377-588-Fc protein efficiently binds to dipeptidyl peptidase 4 (DPP4), the receptor of MERS-CoV, and potently inhibited MERS-CoV infection, suggesting its potential to be further developed as a therapeutic modality for treating MERS-CoV infection and saving the patients' lives. The recombinant S377-588-Fc is able to induce in the vaccinated mice strong MERS-CoV S-specific antibodies, which blocks the binding of RBD to DPP4 receptor and effectively neutralizes MERS-CoV infection. These findings indicate that this truncated RBD protein shows promise for further development as an effective and safe vaccine for the prevention of MERS-CoV infection.

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Construction and characterization of MERS-CoV S377-588-Fc.(A) Schematic structure of MERS-CoV S1 subunit and S377-588-Fc. RBM: the receptor-binding motif in the RBD. S377-588-Fc was constructed by fusing MERS-CoV residues 377-588 of S1 with Fc of human IgG. (B) SDS-PAGE and Western blot (WB) analysis of purified 377-588-Fc protein. Samples were either boiled for 10 min, or not boiled, followed by SDS-PAGE (left) and WB (right) analysis using a S1-specific polyclonal antibody. (C) Analysis of S377-588-Fc protein conformation by cross-linker. Samples were cross-linked with glutaraldehyde (with cross-linker at the final concentration of 4 µM) or without cross-linker (w/o cross-linker), followed by SDS-PAGE (left) and WB (right) analysis as described above. The protein molecular weight marker (kDa) (Invitrogen) is indicated on the left.
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pone-0081587-g001: Construction and characterization of MERS-CoV S377-588-Fc.(A) Schematic structure of MERS-CoV S1 subunit and S377-588-Fc. RBM: the receptor-binding motif in the RBD. S377-588-Fc was constructed by fusing MERS-CoV residues 377-588 of S1 with Fc of human IgG. (B) SDS-PAGE and Western blot (WB) analysis of purified 377-588-Fc protein. Samples were either boiled for 10 min, or not boiled, followed by SDS-PAGE (left) and WB (right) analysis using a S1-specific polyclonal antibody. (C) Analysis of S377-588-Fc protein conformation by cross-linker. Samples were cross-linked with glutaraldehyde (with cross-linker at the final concentration of 4 µM) or without cross-linker (w/o cross-linker), followed by SDS-PAGE (left) and WB (right) analysis as described above. The protein molecular weight marker (kDa) (Invitrogen) is indicated on the left.

Mentions: Unlike SARS-CoV, which uses human angiotensin-converting enzyme 2 (ACE2) as its receptor for binding to ACE2-expressing cells [11], MERS-CoV utilizes a different receptor, dipeptidyl peptidase 4 (DPP4), for binding to DPP4-expressing cells [12]. Like the spike (S) protein of SARS-CoV, the S protein of MERS-CoV also plays important roles in virus entry and infection [13]. MERS-CoV S protein contains a S1 subunit that mediates virus binding to cells expressing DPP4 through its receptor-binding domain (RBD) region and an S2 subunit that mediates virus-cell membrane fusion [12], [13]. Based on sequence alignment and homology modeling analysis and functional studies, we and Mou et al. have predicted that the RBD is located in residues 377-662 or 358-588 of the MERS-CoV S1 subunit [14]–[16] (Fig. 1A). Co-crystallographic analyses of the RBD/DPP4 complexes have confirmed that the RBD is attributed to residues 367-606 or 367-588 in MERS-CoV S1 [17]–[19] (Fig. 1A).


A truncated receptor-binding domain of MERS-CoV spike protein potently inhibits MERS-CoV infection and induces strong neutralizing antibody responses: implication for developing therapeutics and vaccines.

Du L, Kou Z, Ma C, Tao X, Wang L, Zhao G, Chen Y, Yu F, Tseng CT, Zhou Y, Jiang S - PLoS ONE (2013)

Construction and characterization of MERS-CoV S377-588-Fc.(A) Schematic structure of MERS-CoV S1 subunit and S377-588-Fc. RBM: the receptor-binding motif in the RBD. S377-588-Fc was constructed by fusing MERS-CoV residues 377-588 of S1 with Fc of human IgG. (B) SDS-PAGE and Western blot (WB) analysis of purified 377-588-Fc protein. Samples were either boiled for 10 min, or not boiled, followed by SDS-PAGE (left) and WB (right) analysis using a S1-specific polyclonal antibody. (C) Analysis of S377-588-Fc protein conformation by cross-linker. Samples were cross-linked with glutaraldehyde (with cross-linker at the final concentration of 4 µM) or without cross-linker (w/o cross-linker), followed by SDS-PAGE (left) and WB (right) analysis as described above. The protein molecular weight marker (kDa) (Invitrogen) is indicated on the left.
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC3852489&req=5

pone-0081587-g001: Construction and characterization of MERS-CoV S377-588-Fc.(A) Schematic structure of MERS-CoV S1 subunit and S377-588-Fc. RBM: the receptor-binding motif in the RBD. S377-588-Fc was constructed by fusing MERS-CoV residues 377-588 of S1 with Fc of human IgG. (B) SDS-PAGE and Western blot (WB) analysis of purified 377-588-Fc protein. Samples were either boiled for 10 min, or not boiled, followed by SDS-PAGE (left) and WB (right) analysis using a S1-specific polyclonal antibody. (C) Analysis of S377-588-Fc protein conformation by cross-linker. Samples were cross-linked with glutaraldehyde (with cross-linker at the final concentration of 4 µM) or without cross-linker (w/o cross-linker), followed by SDS-PAGE (left) and WB (right) analysis as described above. The protein molecular weight marker (kDa) (Invitrogen) is indicated on the left.
Mentions: Unlike SARS-CoV, which uses human angiotensin-converting enzyme 2 (ACE2) as its receptor for binding to ACE2-expressing cells [11], MERS-CoV utilizes a different receptor, dipeptidyl peptidase 4 (DPP4), for binding to DPP4-expressing cells [12]. Like the spike (S) protein of SARS-CoV, the S protein of MERS-CoV also plays important roles in virus entry and infection [13]. MERS-CoV S protein contains a S1 subunit that mediates virus binding to cells expressing DPP4 through its receptor-binding domain (RBD) region and an S2 subunit that mediates virus-cell membrane fusion [12], [13]. Based on sequence alignment and homology modeling analysis and functional studies, we and Mou et al. have predicted that the RBD is located in residues 377-662 or 358-588 of the MERS-CoV S1 subunit [14]–[16] (Fig. 1A). Co-crystallographic analyses of the RBD/DPP4 complexes have confirmed that the RBD is attributed to residues 367-606 or 367-588 in MERS-CoV S1 [17]–[19] (Fig. 1A).

Bottom Line: Development of effective therapeutics and vaccines is crucial to save lives and halt the spread of MERS-CoV.The recombinant S377-588-Fc is able to induce in the vaccinated mice strong MERS-CoV S-specific antibodies, which blocks the binding of RBD to DPP4 receptor and effectively neutralizes MERS-CoV infection.These findings indicate that this truncated RBD protein shows promise for further development as an effective and safe vaccine for the prevention of MERS-CoV infection.

View Article: PubMed Central - PubMed

Affiliation: Lindsley F. Kimball Research Institute, New York Blood Center, New York, New York, United States of America.

ABSTRACT
An emerging respiratory infectious disease with high mortality, Middle East respiratory syndrome (MERS), is caused by a novel coronavirus (MERS-CoV). It was first reported in 2012 in Saudi Arabia and has now spread to eight countries. Development of effective therapeutics and vaccines is crucial to save lives and halt the spread of MERS-CoV. Here, we show that a recombinant protein containing a 212-amino acid fragment (residues 377-588) in the truncated receptor-binding domain (RBD: residues 367-606) of MERS-CoV spike (S) protein fused with human IgG Fc fragment (S377-588-Fc) is highly expressed in the culture supernatant of transfected 293T cells. The purified S377-588-Fc protein efficiently binds to dipeptidyl peptidase 4 (DPP4), the receptor of MERS-CoV, and potently inhibited MERS-CoV infection, suggesting its potential to be further developed as a therapeutic modality for treating MERS-CoV infection and saving the patients' lives. The recombinant S377-588-Fc is able to induce in the vaccinated mice strong MERS-CoV S-specific antibodies, which blocks the binding of RBD to DPP4 receptor and effectively neutralizes MERS-CoV infection. These findings indicate that this truncated RBD protein shows promise for further development as an effective and safe vaccine for the prevention of MERS-CoV infection.

Show MeSH
Related in: MedlinePlus