Atomic Structures of Minor Proteins VI and VII in Human Adenovirus

Abstract

Human adenoviruses (Ad) are dsDNA viruses associated with infectious diseases, yet better known as tools for gene delivery and oncolytic anti-cancer therapy. Atomic structures of Ad provide the basis for the development of antivirals and for engineering efforts towards more effective applications. Since 2010, atomic models of human Ad5 have been independently derived from photographic film cryoEM and X-ray crystallography, but discrepancies exist concerning the assignment of cement proteins IIIa, VIII and IX. To clarify these discrepancies, here we have employed the technology of direct electron-counting to obtain a cryoEM structure of human Ad5 at 3.2 Å resolution. Our improved structure unambiguously confirmed our previous cryoEM models of proteins IIIa, VIII and IX and explained the likely cause of conflict in the crystallography models. The improved structure also allows the identification of three new components in the cavities of hexons - the cleaved N-terminus of precursor protein VI (pVIn), the cleaved N-terminus of precursor protein VII (pVIIn2), and mature protein VI. The binding of pVIIn2--by extension that of genome-condensing pVII--to hexons is consistent with the previously proposed dsDNA genome-capsid co-assembly for adenoviruses, which resembles that of ssRNA viruses but differs from the well-established mechanism of pumping dsDNA into a preformed protein capsid, as exemplified by tailed bacteriophages and herpesviruses. Adenovirus is a double-edged sword to humans - as a widespread pathogen and a bioengineering tool for anti-cancer and gene therapy. Atomic structure of the virus provides the basis for antiviral and application developments, but conflicting atomic models from conventional/film cryoEM and X-ray crystallography for important cement proteins IIIa, VIII, and IX have caused confusion. Using the cutting-edge cryoEM technology with electron counting, we improved the structure of human adenovirus type 5 and confirmed our previous models of cement proteins IIIa, VIII, and IX, thus clarifying the inconsistent structures. The improved structure also reveals atomic details of membrane-lytic protein VI and genome-condensing protein VII and supports the previously proposed genome-capsid co-assembly mechanism for adenoviruses.

Dynamic competition for hexon binding between core protein VII and lytic protein VI promotes adenovirus maturation and entry

“…The second study revealing unexpected relationships between proteins VI and VII is the most recent cryo-electron microscopy (cryo-EM) analysis of the HAdV-C5 capsid structure (2).…”

“…First, at the mouth of the hexon cavity, residues 109 to 143 of pVI were modeled, disconnected from pVI N . Second, the pVI N tracing was reversed with respect to the previous structures (31,32), in such a way that in the latest model (2) the pVI N cleavage site is not accessible at the rim of the hexon cavity, but hidden inside and oriented away from the core. This new disposition makes it more difficult to understand how AVP, sliding on the DNA, can reach its target sequence in pVI N (Fig.…”

Bovine adenovirus‐3 protein VIII associates with eukaryotic initiation factor‐6 during infection

“…Although these studies incredibly advanced the understanding of adenovirus structure, they also created some confusion regarding the location of important minor proteins. Although all the previous studies reported that two copies of protein VIII are present on the inner side of the capsid, there were discrepancies regarding surrounding minor proteins (Dai, Wu, Sun, & Zhou, 2017). Although one study reported that protein IIIa and protein VIII are present at inner surface of capsid around each fivefold axis (Liu et al, 2010), another study reported that instead of protein IIIa, proteins V and VI are present at fivefold axis and form a ternary complex with protein VIII.…”

Epigenetics and the dynamics of chromatin during adenovirus infections

“…The interaction of proteins within the adenovirus core was studied in the 1970s and 1980s [27][28][29][32][33][34][35][36], establishing that protein VII directly interacts with viral DNA. Recent structural studies propose that protein VII is integral to the virion assembly process [37]; however, the formation of adenovirus particles in the complete absence of protein VII [38] implies that the viral genome conformation may have multiple states that are amenable to virion formation. Recent structural studies propose that protein VII is integral to the virion assembly process [37]; however, the formation of adenovirus particles in the complete absence of protein VII [38] implies that the viral genome conformation may have multiple states that are amenable to virion formation.…”

“…Through interactions with protein V, protein VII also stabilizes the genome within the capsid. Recent structural studies propose that protein VII is integral to the virion assembly process [37]; however, the formation of adenovirus particles in the complete absence of protein VII [38] implies that the viral genome conformation may have multiple states that are amenable to virion formation. It has been suggested recently that one conformation may resemble a more ancient structure similar to archaeal chromatin [39,40], although without further structural insight into protein VII-DNA complexes, the conformation of viral genomes within virions remains unclear.…”