Modest truncation of the major capsid protein abrogates B19 parvovirus capsid formation

Canine parvovirus capsids are composed of 60 copies of VP2 and 6 to 10 copies of VP1. To locate essential sites of interaction between VP2 monomers, we have analyzed the effects of a number of VP2 deletion mutants representing the amino terminus and the four major loops of the surface, using as an assay the formation of virus-like particles (VLPs) expressed by recombinant baculoviruses. For the amino terminus we constructed three mutants with progressively larger deletions, i.e., 9, 14, and 24 amino acids. Deletions of 9 and 14 amino acids did not affect the morphology and assembly capabilities of the mutants. However, the mutant with the 24-amino-acid deletion did not show hemagglutination properties or correct VLP morphology, stressing again the relevance of the RNER domain in canine parvovirus functionality. Three of the four mutants with deletions in the loops failed to make correct VLPs, indicating that these regions are essential for correct capsid assembly and morphology. Only the mutant with the deletion in loop 2 was able to assemble in regular VLPs, suggesting that this loop has little or no effect in capsid morphogenesis. Further research has demonstrated that this region can tolerate the insertion of foreign epitopes that are correctly exposed in the surface of the capsid. This result opens the door to the use of these VLPs for antigen delivery.

Section: Discussioncontrasting

confidence: 99%

“…The disorder of the amino termini of CPV VP2 was assumed to be due to association of this region with the viral DNA (10). For this reason, no participation of the amino-terminal domain in capsid conformation and morphogenesis was expected.…”

Section: Discussionmentioning

confidence: 99%

Identification of domains in canine parvovirus VP2 essential for the assembly of virus-like particles

Hurtado

Rueda

Nowicky

et al. 1996

“…However, it is also possible that other mutations within the A. sylvaticus or R. norvegicus EVE sequences or our other repair sequences prevented assembly. Studies with extant parvoviruses show that single point mutations within the major capsid protein can block capsid assembly, especially when they occur within the well-conserved residues that form the interior of the capsid or are near the 5-fold axis (36)(37)(38)(39). Expressing many of the less conserved exterior surface loops in place of the equivalent sequences of the CPV capsid allowed the chimeras to assemble (Fig.…”

Section: Discussionmentioning

confidence: 99%

Examination and Reconstruction of Three Ancient Endogenous Parvovirus Capsid Protein Gene Remnants Found in Rodent Genomes

Callaway

Subramanian

Urbina

et al. 2019

Parvovirus-derived endogenous viral elements (EVEs) have been found in the genomes of many different animal species, resulting from integration events that may have occurred from more than 50 million years ago to much more recently. Here, we further investigate the properties of autonomous parvovirus EVEs and describe their relationships to contemporary viruses. While we did not find any intact capsid protein open reading frames in the integrated viral sequences, we examined three EVEs that were repaired to form full-length sequences with relatively few changes. These sequences were found in the genomes of Rattus norvegicus (brown rat), Mus spretus (Algerian mouse), and Apodemus sylvaticus (wood mouse). The R. norvegicus sequence was not present in the genomes of the closely related species R. rattus, R. tanezumi, R. exulans, and R. everetti, indicating that it was less than 2 million years old, and the M. spretus and A. sylvaticus sequences were not found in the published genomes of other mouse species, also indicating relatively recent insertions. The M. spretus VP2 sequence assembled into capsids, which had high thermal stability, bound the sialic acid N-acetylneuraminic acid, and entered murine L cells. The 3.89-Å structure of the M. spretus virus-like particles (VLPs), determined using cryo-electron microscopy, showed similarities to rodent and porcine parvovirus capsids. The repaired VP2 sequences from R. norvegicus and A. sylvaticus did not assemble as first prepared, but chimeras combining capsid surface loops from R. norvegicus with canine parvovirus assembled, allowing some of that capsid’s structures and functions to be examined. IMPORTANCE Parvovirus endogenous viral elements (EVEs) that have been incorporated into the genomes of different animals represent remnants of the DNA sequences of ancient viruses that infected the ancestors of those animals millions of years ago, but we know little about their properties or how they differ from currently circulating parvoviruses. By expressing the capsid proteins of different parvovirus EVEs that were found integrated into the genomes of three different rodents, we can examine their structures and functions. A VP2 (major capsid protein) EVE sequence from a mouse genome assembled into capsids that had a similar structure and biophysical properties to extant parvoviruses and also bound sialic acids and entered rodent cells. Chimeras formed from combinations of canine parvovirus and portions of the parvovirus sequences from the brown rat genome allowed us to examine the structures and functions of the surface loops of that EVE capsid.

“…In contrast to AAV, it has been shown that the capsids of these parvoviruses can be formed in the presence of the major capsid protein, VP2, alone (4,9). In previous work, it was shown that the most N-terminal 25amino-acid residues of VP2 of B19 were not required for capsid assembly (10). Amino acid sequence alignment of VP2 protein of B19 with AAV indicates that the minimal protein capable of B19 capsid formation corresponds to VP3 rather than VP2 of AAV (data not shown).…”

mentioning

confidence: 99%

Nuclear Transport of the Major Capsid Protein Is Essential for Adeno-Associated Virus Capsid Formation

Hoque

Ishizu

Matsumoto

et al. 1999

Adeno-associated virus capsids are composed of three proteins, VP1, VP2, and VP3. Although VP1 is necessary for viral infection, it is not essential for capsid formation. The other capsid proteins, VP2 and VP3, are sufficient for capsid formation, but the functional roles of each protein are still not well understood. By analyzing a series of deletion mutants of VP2, we identified a region necessary for nuclear transfer of VP2 and found that the efficiency of nuclear localization of the capsid proteins and the efficiency of virus-like particle (VLP) formation correlated well. To confirm the importance of the nuclear localization of the capsid proteins, we fused the nuclear localization signal of simian virus 40 large T antigen to VP3 protein. We show that this fusion protein could form VLP, indicating that the VP2-specific region located on the N-terminal side of the protein is not structurally required. This finding suggests that VP3 has sufficient information for VLP formation and that VP2 is necessary only for nuclear transfer of the capsid proteins.