JOURNAL OF VIROLOGY, June 2010, p. 5880–5889 Vol. 84, No. 120022-538X/10/$12.00 doi:10.1128/JVI.02719-09Copyright © 2010, American Society for Microbiology. All Rights Reserved.Human Bocavirus Capsid Structure: Insights into the StructuralRepertoire of the Parvoviridae䌤Brittney L. Gurda,1† Kristin N. Parent,2† Heather Bladek,1Robert S. Sinkovits,2Michael A. DiMattia,1Chelsea Rence,1Alejandro Castro,1Robert McKenna,1Norm Olson,2Kevin Brown,4Timothy S. Baker,2,3* and Mavis Agbandje-McKenna1*Department of Biochemistry and Molecular Biology, University of Florida, Gainesville, Florida 326101; Department of Chemistryand Biochemistry2and Division of Biological Sciences,3University of California, San Diego, California 92093; andVirus Reference Department, Centre for Infections, Health Protection Agency, London NW9 5HT, United Kingdom4Received 26 December 2009/Accepted 26 March 2010Human bocavirus (HBoV) was recently discovered and classified in the Bocavirus genus (family Parvoviridae,subfamily Parvovirinae) on the basis of genomic similarity to bovine parvovirus and canine minute virus. HBoVhas been implicated in respiratory tract infections and gastroenteric disease in children worldwide, yet despitenumerous epidemiological reports, there has been limited biochemical and molecular characterization of thevirus. Reported here is the three-dimensional structure of recombinant HBoV capsids, assembled from viralprotein 2 (VP2), at 7.9-Å resolution as determined by cryo-electron microscopy and image reconstruction. Apseudo-atomic model of HBoV VP2 was derived from sequence alignment analysis and knowledge of the crystalstructure of human parvovirus B19 (genus Erythrovirus). Comparison of the HBoV capsid structure to that ofparvoviruses from five separate genera demonstrates strong conservation of a ␤-barrel core domain and an␣-helix, from which emanate several loops of various lengths and conformations, yielding a unique surfacetopology that differs from the three already described for this family. The highly conserved core is consistentwith observations for other single-stranded DNA viruses, and variable surface loops have been shown to conferthe host-specific tropism and the diverse antigenic properties of this family.Human bocavirus (HBoV), a newly discovered member ofthe family Parvoviridae, was originally isolated in randomlyselected nasopharyngeal aspirates (5). Since this initial discov-ery, HBoV has also been detected worldwide, predominantlyin children under the age of 2 years with respiratory infections,in serum, urine, and fecal samples (40). Symptomatic childrencommonly exhibit acute diseases of the upper and lower respi-ratory tracts (7, 36, 44, 56) and, possibly, gastroenteritis (31,56) though the link to gastroenteritis outbreaks has been ques-tioned (12). It is still unclear if HBoV is the sole etiologic agentof respiratory disease as higher rates of coinfections with otherrespiratory pathogens such as human rhinovirus and Strepto-coccus spp. are often observed (4). However, Allander et al.recently reported (4) that HBoV was found in 19% of childrenwith acute wheezing, thereby making it the fourth most com-mon virus, after rhinoviruses, enteroviruses, and respiratorysyncytial virus, detected in children exhibiting this symptom.These findings suggest that, at high viral load, HBoV could bean etiologic agent of respiratory tract disease (4). HBoV in-fection is common in the first few years of life, and clinicalresearch suggests it may follow the primary period for acqui-sition of human parvovirus B19 (B19) though there is no an-tigenic cross-reactivity between B19 and HBoV (28, 30). Byage 5, most people have circulating antibodies against HBoV,as is also true for other respiratory viruses such as respiratorysyncytial virus, rhinoviruses, and human metapneumovirus(17). HBoV has also been identified in adults, with ⬃63% ofsamples tested being seropositive, showing a positive correla-tion with age and a slight positive bias toward women (14).The Parvoviridae is a family of small, nonenveloped virusesthat package a single-stranded DNA (ssDNA) genome of⬃5,000 bases. These viruses are subdivided into two subfami-lies: Parvovirinae and Densovirinae (Table 1). The Parvovirinaeare further subdivided into five genera, all of whose membersinfect vertebrates. The Densovirinae (four genera) infect onlyinvertebrates. Phylogenetic analysis places HBoV in the re-cently classified Bocavirus genus (Table 1). In addition toHBoV, numerous parvoviruses circulate among the humanpopulation. Among these are the following: several dependovi-ruses; adeno-associated virus (AAV) serotypes AAV1 toAAV3, AAV5, and AAV9; the Erythrovirus B19; and the newlydiscovered human parvovirus genotypes 4 (Parv4) and 5(Parv5) (23, 27, 50). Of these, only B19 had been implicated indisease until the discovery of HBoV and Parv4, which has beenisolated from patients who present symptoms of acute HIVinfection (50).The HBoV genome, like that of all members of the Bocavi-rus genus, contains three open reading frames (ORFs). Thefirst ORF, at the 5⬘ end, encodes NS1, a nonstructural protein.The next ORF, unique to the bocaviruses, encodes NP1, asecond nonstructural protein. The third ORF, at the 3⬘ end,encodes the two structural capsid viral proteins (VPs), VP1and VP2. The HBoV VPs share 42% and 43% amino acid* Corresponding author. Mailing address for T. S. Baker: Depart-ment of Chemistry and Biochemistry, University of California, SanDiego, CA 92093. Phone: (858) 534-5845. Fax: (858) 534-5846. E-mail:[email protected]. Mailing address for M. Agbandje-McKenna:Department of Biochemistry and Molecular Biology, University ofFlorida, P.O. Box 100245, Gainesville, FL 32610. Phone: (352) 3925694. Fax: (352) 392-3422. E-mail: [email protected].† B.L.G. and K.N.P. contributed equally to this work.䌤Published ahead of print on 7 April 2010.5880 at UNIV OF CALIFORNIA SAN DIEGO on May 18, 2010 jvi.asm.orgDownloaded fromsequence identity with the corresponding VPs of bovine par-vovirus and canine minute virus, respectively (5). More re-cently, two additional HBoV-like viruses, HBoV-2 andHBoV-3, were identified in stool samples from children (8, 31).The genome organization of these viruses is identical to that ofHBoV, with the NS1, NP1, and VP proteins of HBoV-2 andHBoV-3 being, respectively, ⬃80 and 90%, ⬃70 and 80%, and⬃80 and 80% identical to the respective proteins in HBoV (8,31).Parvovirus genomes are