General Background

Adeno-associated viruses (AAVs) are members of the Parvoviridae family.The AAVs are distinct from the autonomous parvoviruses by their dependence on a helper virus for replication. These viruses require coinfection with either an adenovirus or a herpesvirus for replicative infection. To date, nine distinct serotypes, AAV1-5, AAV7-9, and AAAV, have been cloned from either human or primate samples. In addition, a large number of sequences were recently isolated from monkeys. There are no diseases associated with AAVs, and their capability of packaging foreign DNA makes them attractive for development as gene therapy vectors.

Structure

The AAV virions have a T = 1 icosahedral capsid consisting of 60 copies of three related proteins, VP1, VP2, and VP3, at an estimated ratio of 1:1:8, which surrounds a singlestranded DNA genome. These three proteins share a common C-terminal region (the ~590 amino acids of VP3) but have different amino termini resulting from alternative start codon usage. The entire sequence of VP3 is present in VP2, whose sequence is in turn entirely contained within VP1. The VP3 common region appears to be essential for cell binding and antigenic properties and possibly plays a role in genomic DNA packaging. VP1 of AAV serotype 2 (AAV2) has a unique N-terminal region of ~130 amino acids, which is important in the viral life cycle after cell binding and entry, and displays a phospholipase A2 function that also appears to be conserved in other serotypes.

The three-dimensional (3D) structures of several autonomous parvoviruses and that of AAV2 have been determined by X-ray crystallography. Lower resolution structures of AAV2 and AAV5, as well as those of the autonomous parvoviruses Aleutian mink disease parvovirus (ADV) and human parvovirus B19, have been mapped by a combination of cryo-electron microscopy (cryo-EM) and pseudoatomic model building. In all of these structures, only the C-terminal common region of VP1/2/3 is observed. The core of the protein is composed of a conserved eight-stranded antiparallel b-barrel motif. The majority of the variable surface structure consists of large loops inserted between the strands of the b-barrel. Structural features on the capsid surfaces of these viruses include projections at or surrounding the icosahedral threefold axis and depressions at the twofold and around the fivefold axes. A conserved cylindrical channel is present at the icosahedral fivefold axis formed by symmetry-related b-ribbons.

Molecular Characterization

Biochemical and molecular characterizations of several of the different serotypes of AAV have indicated that each has unique cell binding characteristics and tissue transduction efficiencies. AAV2, the best characterized serotype, has a broad tropism, including tropisms for the eye, central nervous system (CNS), liver, and muscle, and has been studied for the treatment of genetic diseases such as cystic fibrosis and alpha 1 antitrypsin deficiency. However, vectors derived from other AAV serotypes have been reported to be more efficient at transducing certain cell types than AAV2. AAV1 has demonstrated an improved tropism for striated muscle cells compared to AAV2. AAV3 can transduce hematopoietic cells. AAV4, originally isolated from African green monkeys, is able to infect cells from humans and rodents. The direct injection of AAV4 into the striata of mice demonstrated a strong tropism for ependymal cells in the CNS. In vitro and in vivo experiments with AAV5 have demonstrated improved binding and transduction of airway lung epithelia, muscles, CNS neurons, and the eye compared to AAV2. The more recently identified AAV serotypes, including AAV7 and AAV8, also appear to have unique cell tropisms. For example, AAV8 is reported to efficiently transduce liver cells. These differences in cell tropism appear to be due to the ability of the AAV capsids to utilize different cell surface carbohydrates and/or protein receptors for cell binding and entry.

Our Focus

Our lab has many projects in conjunction with the Muzyczka Lab that involves all aspects of AAV capsid structure, and its role in the virus life cycle. We are interested in using various structural techniques (from X-ray crystallography, cryo-EM to molecular modeling) to study the detailed structures of wild type and various mutant of AAVs.

Selected Recent Publications

Padron, E., V. Bowman, N, Kaludov, L. Govindasamy, H. Levy, P. Nick, R. McKenna, N. Muzyczka, J. A. Chiorini, T. S. Baker, M. Agbandje-McKenna. 2005. The structure of adeno-associated virus type4. J. Virol., 79:5047-5058.
Padron 2005

Opie, S.R., K.H. Warrington Jr, M. Agbandje-McKenna, S. Zolotukhin, N. Muzyczka.2003. Identification of amino acid residues in the capsid proteins of adeno-associated virus type 2 that contribute to heparan sulfate proteoglycan binding. J. Virology, 77:6995-7006.
Opie 2003