Adeno-Associated Virus (AAV-DJ)—Cryo-EM Structure at 1.56 Å Resolution

Xie, Qing; Yoshioka, Craig; Chapman, Michael S.

doi:10.3390/v12101194

Cited by 21 publications

(31 citation statements)

References 64 publications

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“…OPEN ACCESS resolution structures for complexes recalcitrant to crystallization, but now cryo-EM regularly equals or exceeds X-ray resolutions [60]. The large size and symmetry of viral assemblies renders them particularly amenable.…”

Section: Trends In Microbiologymentioning

confidence: 99%

“…The large size and symmetry of viral assemblies renders them particularly amenable. Occasionally, exceptional resolution is achieved, with AAV-DJ, at 1.56 Å, among the highest resolution biomolecular cryo-EM structures [60]. Cryo-EM is now also providing a path towards atomic structures at crystallographiclike resolutions of complexes with protein receptors that would be difficult or impossible to crystallize.…”

Section: Trends In Microbiologymentioning

confidence: 99%

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Adeno-associated virus (AAV) cell entry: structural insights

Meyer

Chapman

2022

Trends in Microbiology

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Adeno-associated virus (AAV) is the leading vector in emerging treatments of inherited diseases. Higher transduction efficiencies and cellular specificity are required for broader clinical application, motivating investigations of virus-host molecular interactions during cell entry. High-throughput methods are identifying host proteins more comprehensively, with subsequent molecular studies revealing unanticipated complexity and serotype specificity. Cryogenic electron microscopy (cryo-EM) provides a path towards structural details of these sometimes heterogeneous virus-host complexes, and is poised to illuminate more fully the steps in entry. Here presented, is progress in understanding the distinct steps of glycan attachment, and receptor-mediated entry/trafficking. Comparison with structures of antibody complexes provides new insights on immune neutralization with implications for the design of improved gene therapy vectors.Virus-host interactions: a foundation for improved gene therapy vectors AAV (see Glossary) is a small, single-stranded DNA virus of the family Parvoviridae whose three viral capsid proteins (VP1, VP2, and VP3) comprise a non-enveloped shell of icosahedral symmetry [1]. Since wild-type (wt) AAVs possess wide tissue tropism and low pathogenicity (as replication-deficient dependoparvoviruses), recombinantly produced AAV vectors (rAAVs) are now pre-eminent vehicles for therapeutic gene delivery, with clinical payloads replacing the ~4.7 kb native genome. The past 3 years have seen FDA approval of two AAV therapeutics, LUXTURNA™ (rAAV2 for RPE65-related retinal dystrophy) and ZOLGENSMA® (rAAV9 for spinal muscular atrophy). Despite successes, target-specific vector optimization and higher transduction efficiency remain crucial for wider clinical application [2]. We find that each AAV capsid variant discovered, engineered, or derived, can reveal, through its unique sequence-encoded phenotype, clues to improving targeting, transduction, and immune evasion. AAV capsid interactions with host cell entry factors, in particular, have been a focus of recent research, with exciting results.Advances in high-throughput screening and structure determination are being employed by the AAV community to identify and visualize AAV-host molecule interactions critical to cell transduction. Increasingly high-resolution cryo-EM structures help to elucidate the molecular determinants of receptor, antibody, and cell-surface glycan interactions. Here we summarize the current understanding of AAV entry factors and their identification, and the potential for high-resolution structures to inform future vector design. A recent review by Riyad and Weber [3] presents an excellent perspective on the cellular processes of AAV entry and trafficking, while, here, emphasis is on the biophysical chemistry and detailed characterization of molecular interactions. AAV cell entry: sequence of eventsPromiscuous attachment of AAVs to glycans on the cell surface is thought to increase the likelihood of internalization through increased en...

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Section: Trends In Microbiologymentioning

confidence: 99%

Section: Trends In Microbiologymentioning

confidence: 99%

Adeno-associated virus (AAV) cell entry: structural insights

Meyer

Chapman

2022

Trends in Microbiology

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“…Combining the high-resolution apostructures of AAV with extensive mutational data, an overview of the binding site locations and their residues has been obtained. A recent cryo-EM structure of AAV-DJ at a resolution below 1.6 Å [ 21 ], and a continuing interest in AAV as a gene therapy vector [ 22 ], shows this is an active field, both in structural biology and applied medicine; therefore, elucidating the details of AAV–GAG interactions at atomic (or near atomic) resolution would be an important development.…”

Section: Adeno-associated Virusmentioning

confidence: 99%

Structural Insight into Non-Enveloped Virus Binding to Glycosaminoglycan Receptors: A Review

Sorin

Kuhn

Stasiak

et al. 2021

Viruses

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Viruses are infectious agents that hijack the host cell machinery in order to replicate and generate progeny. Viral infection is initiated by attachment to host cell receptors, and typical viral receptors are cell-surface-borne molecules such as proteins or glycan structures. Sialylated glycans (glycans bearing sialic acids) and glycosaminoglycans (GAGs) represent major classes of carbohydrate receptors and have been implicated in facilitating viral entry for many viruses. As interactions between viruses and sialic acids have been extensively reviewed in the past, this review provides an overview of the current state of structural knowledge about interactions between non-enveloped human viruses and GAGs. We focus here on adeno-associated viruses, human papilloma viruses (HPVs), and polyomaviruses, as at least some structural information about the interactions of these viruses with GAGs is available. We also discuss the multivalent potential for GAG binding, highlighting the importance of charged interactions and positively charged amino acids at the binding sites, and point out challenges that remain in the field.

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“…In particular, cryo-TEM has recently achieved substantially improved resolution due to the incorporation of highsensitivity cameras. Its resolution approaches atomic resolution, similar to X-ray crystallography, despite dealing with dispersed protein particles [1][2][3][4][5] . This cryo-TEM method developed under such circumstances is naturally targeted to the structural analysis of protein molecules.…”

Section: Introductionmentioning

confidence: 99%

Development of a New cryo-S(T)EM Technique Allowing Simultaneous STEM and SEM Imaging and its Application to Biological Samples

Usukura¹,

Narita²,

Matsumoto³

et al. 2021

Preprint

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A new type of cryo-electron microscopy (cryo-S(T)EM) technique made possible by installing a new cryo-transfer holder and an anti-contamination trap on a scanning electron microscope (Hitachi SU9000) allowed simultaneous collection of both transmission (transmission electron microscopy, TEM) images and surface (scanning electron microscopy, SEM) images at -180°C. The ultimate temperatures of the cryo-transfer holder and the anti-contamination trap reached − 190°C and − 210°C, respectively, by applying a liquid nitrogen slush. The TEM images obtained by the new cryo-S(T)EM method showed quality equal or superior to that of images obtained by conventional 100 kV TEM, although the resolution did not improve at -180°C due to slight drifting of the sample stage. Cryo-S(T)EM also had the unexpected advantage of enabling observations of intracellular structures in thick frozen cells by accelerating the sublimation of ice surrounding the specimens. The spatial architecture of the cytoskeleton, poly-ribosome-chains, endoplasmic reticulum (ER), mitochondria, etc., became visible in thick frozen cells via sufficient (deep) sublimation of ice in combination with the unroofing method. In particular, it should be noted that the ER appeared as a wide and flat structure beneath the cell membrane while forming a large spatial network together with tubular ER.

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Adeno-Associated Virus (AAV-DJ)—Cryo-EM Structure at 1.56 Å Resolution

Cited by 21 publications

References 64 publications

Adeno-associated virus (AAV) cell entry: structural insights

Adeno-associated virus (AAV) cell entry: structural insights

Structural Insight into Non-Enveloped Virus Binding to Glycosaminoglycan Receptors: A Review

Development of a New cryo-S(T)EM Technique Allowing Simultaneous STEM and SEM Imaging and its Application to Biological Samples

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