Structural analyses at pseudo atomic resolution of Chikungunya virus and antibodies show mechanisms of neutralization

Sun, Siyang; Xiang, Ye; Akahata, Wataru; Holdaway, Heather A.; Pal, Pankaj Kumar; Zhang, Xinzheng; Diamond, Michael S.; Nabel, Gary J.; Rossmann, Michael G.

doi:10.7554/elife.00435

Cited by 150 publications

(206 citation statements)

References 86 publications

(110 reference statements)

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“…The most obvious characteristic of the fitting results is that the density of the E2-B domain is lower for the 4J21 Fab complex compared with the cryo-EM density for the other E1 and E2 domains. A comparable reduction of the E2-B domain density was observed for the uncomplexed structure (22). This finding indicates that the B domain has greater flexibility relative to the other domains in the uncomplexed structure.…”

Section: Resultssupporting

confidence: 65%

“…The only previous structural studies of CHIKVantibody complexes were of four neutralizing murine mAbs. Three of these antibodies (m10, m242, and CHK-9) bound to the putative receptor binding sites, whereas one, CHK-152, inhibited the exposure of the fusion loop by immobilizing the E2-B domain (22). Analogous structural studies have been reported for other alphaviruses, such as Ross River virus (27), Sindbis virus (33), and VEEV (34).…”

Section: Significancementioning

confidence: 69%

“…Because both mAbs bind close to the putative receptor attachment site (22,26), they also might affect attachment to cells in addition to inhibiting fusion (Fig. 3), although this requires experimental confirmation and the identification of a bona fide CHIKV receptor.…”

Section: Resultsmentioning

confidence: 99%

“…The structure of E2 also has three domains, E2-A, E2-B, and E2-C, and a β-ribbon connector. The E2-A domain contains the putative receptor binding site (22,26,27). The fusion loop in E1-II is covered by the E2-B domain at neutral pH but becomes exposed for membrane fusion in an acidic environment (25,26).…”

Section: Significancementioning

confidence: 99%

“…A cryo-EM structure of CHIKV VLPs has been determined to 5.3-Å resolution (22). Like other alphaviruses, CHIKV is icosahedral and has T = 4 quasi-symmetry (Fig.…”

mentioning

confidence: 99%

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Cryo-EM structures elucidate neutralizing mechanisms of anti-chikungunya human monoclonal antibodies with therapeutic activity

Long

Fong

Austin

et al. 2015

Proc. Natl. Acad. Sci. U.S.A.

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Chikungunya virus (CHIKV) is a mosquito-transmitted alphavirus that causes severe acute and chronic disease in humans. Although highly inhibitory murine and human monoclonal antibodies (mAbs) have been generated, the structural basis of their neutralizing activity remains poorly characterized. Here, we determined the cryo-EM structures of chikungunya virus-like particles complexed with antibody fragments (Fab) of two highly protective human mAbs, 4J21 and 5M16, that block virus fusion with host membranes. Both mAbs bind primarily to sites within the A and B domains, as well as to the B domain's β-ribbon connector of the viral glycoprotein E2. The footprints of these antibodies on the viral surface were consistent with results from loss-of-binding studies using an alanine scanning mutagenesis-based epitope mapping approach. The Fab fragments stabilized the position of the B domain relative to the virus, particularly for the complex with 5M16. This finding is consistent with a mechanism of neutralization in which anti-CHIKV mAbs that bridge the A and B domains impede movement of the B domain away from the underlying fusion loop on the E1 glycoprotein and therefore block the requisite pH-dependent fusion of viral and host membranes. chikungunya virus-antibody complexes | cryo-electron microscopy structure | neutralizing mechanism | viral fusion inhibition C hikungunya virus (CHIKV) is an enveloped, positive-stranded RNA virus that belongs to the alphavirus genus of the Togaviridae family (1, 2). CHIKV is transmitted to humans by Aedes species mosquitoes and causes a debilitating febrile illness associated with acute and chronic arthritis (3, 4). Since the first human CHIKV infection was reported in East Africa in 1952 (5), epidemics of CHIKV have occurred in Africa, Asia, and Europe (6, 7). A CHIKV outbreak in the Caribbean area in late 2013 spread through the Americas and caused about 1.4 million infections (8). Despite its global disease burden and risk of spread, there is no available vaccine or effective antiviral drug for CHIKV.The genome of CHIKV is ∼11.8 kb long and encodes nine viral proteins, five of which are structural (capsid, E3, E2, 6K, and E1) (2). These structural proteins are translated as a single polyprotein, which is then cleaved into the capsid, p62, 6K, and E1 proteins by cellular and viral proteases. During maturation, p62 is cleaved to release E3, which protects the fusion loop in the immature virus. The virus consists of a central core with diameter of ∼400 Å with the icosahedrally organized capsid proteins surrounding the viral genome. The nucleocapsid core is enveloped by a lipid membrane into which the E1 and E2 glycoproteins are inserted (9). The mature CHIKV particle has a diameter of 700 Å. The E2 glycoprotein binds to uncharacterized cellular receptors to initiate virus entry into host cells, whereas E1 glycoprotein participates in virus-host cell membrane fusion (10, 11). Although the E3 and 6K proteins contribute to virus assembly and maturation, they are released during the format...

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Section: Resultssupporting

confidence: 65%

Section: Significancementioning

confidence: 69%

Section: Resultsmentioning

confidence: 99%

Section: Significancementioning

confidence: 99%

“…A cryo-EM structure of CHIKV VLPs has been determined to 5.3-Å resolution (22). Like other alphaviruses, CHIKV is icosahedral and has T = 4 quasi-symmetry (Fig.…”

mentioning

confidence: 99%

See 3 more Smart Citations

Cryo-EM structures elucidate neutralizing mechanisms of anti-chikungunya human monoclonal antibodies with therapeutic activity

Long

Fong

Austin

et al. 2015

Proc. Natl. Acad. Sci. U.S.A.

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Enveloped virus‐like particles as vaccines against pathogenic arboviruses

Pijlman

2015

Biotechnology Journal

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Arthropod-borne arboviruses form a continuous threat to human and animal health, but few arboviral vaccines are currently available. Advances in expression technology for complex, enveloped virus-like particles (eVLPs) create new opportunities to develop potent vaccines against pathogenic arboviruses. In this short review, I highlight the successes and challenges in eVLP production for members of the three major arbovirus families: Flaviviridae (e.g., dengue, West Nile, Japanese encephalitis); Bunyaviridae (e.g., Rift Valley fever); and Togaviridae (e.g., chikungunya). The results from pre-clinical testing will be discussed as well as specific constraints to the large-scale manufacture and purification of eVLPs, which are complex assemblies of membranes and viral glycoproteins. Insect cells emerge as ideal substrates for correct arboviral glycoprotein folding and posttranslational modification to yield high quality eVLPs. Furthermore, baculovirus expression in insect cell culture is scalable and has a proven safety record in industrial human and veterinary vaccine manufacturing. In conclusion, eVLPs produced in insect cells using modern biotechnology have a realistic potential to be used in novel vaccines against arboviral diseases.

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Single‐residue posttranslational modification sites at the N‐terminus, C‐terminus or in‐between: To be or not to be exposed for enzyme access

et al. 2015

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Many protein posttranslational modifications (PTMs) are the result of an enzymatic reaction. The modifying enzyme has to recognize the substrate protein's sequence motif containing the residue(s) to be modified; thus, the enzyme's catalytic cleft engulfs these residue(s) and the respective sequence environment. This residue accessibility condition principally limits the range where enzymatic PTMs can occur in the protein sequence. Non‐globular, flexible, intrinsically disordered segments or large loops/accessible long side chains should be preferred whereas residues buried in the core of structures should be void of what we call canonical, enzyme‐generated PTMs. We investigate whether PTM sites annotated in UniProtKB (with MOD_RES/LIPID keys) are situated within sequence ranges that can be mapped to known 3D structures. We find that N‐ or C‐termini harbor essentially exclusively canonical PTMs. We also find that the overwhelming majority of all other PTMs are also canonical though, later in the protein's life cycle, the PTM sites can become buried due to complex formation. Among the remaining cases, some can be explained (i) with autocatalysis, (ii) with modification before folding or after temporary unfolding, or (iii) as products of interaction with small, diffusible reactants. Others require further research how these PTMs are mechanistically generated in vivo.

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Structural analyses at pseudo atomic resolution of Chikungunya virus and antibodies show mechanisms of neutralization

Cited by 150 publications

References 86 publications

Cryo-EM structures elucidate neutralizing mechanisms of anti-chikungunya human monoclonal antibodies with therapeutic activity

Cryo-EM structures elucidate neutralizing mechanisms of anti-chikungunya human monoclonal antibodies with therapeutic activity

Enveloped virus‐like particles as vaccines against pathogenic arboviruses

Single‐residue posttranslational modification sites at the N‐terminus, C‐terminus or in‐between: To be or not to be exposed for enzyme access

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