Penton Proteins Research Articles

3PO, a novel nonviral gene delivery system using engineered Ad5 penton proteins.

This study describes the development of 3PO, a nonviral, protein-based gene delivery vector which utilizes the highly evolved cell-binding, cell-entry and intracellular transport functions of the adenovirus serotype 5 (Ad5) capsid penton protein. A penton fusion protein containing a polylysine sequence was produced by recombinant methods and tested for gene delivery capability. As the protein itself is known to bind integrins through a conserved consensus motif, the penton inherently possesses the ability to bind and enter cells through receptor-mediated internalization. The ability to lyse the cellular endosome encapsulating internalized receptors is also attributed to the penton. The recombinant protein gains the additional function of DNA binding and transport with the appendage of a polylysine motif. This protein retains the ability to form pentamers and mediates delivery of a reporter gene to cultured cells. Interference by oligopeptides bearing the integrin binding motif suggests that delivery is mediated specifically through integrin receptor binding and internalization. The addition of protamine to penton-DNA complexes allows gene delivery in the presence of serum.

Identification, cloning and sequence analysis of the equine adenovirus 1 hexon gene.

Based on sequence homology with human adenovirus 2 (HAdV2), the hexon gene of equine adenovirus 1 (EAdV1) was identified. HindIII restriction fragments containing the hexon and other viral genes were cloned into the plasmids pUC19 and pBlueScript SK(-) and sequenced. The nucleotide sequence of the hexon gene was completely determined and partial sequence data were obtained for seven other EAdV1 genes. Amino acid (aa) sequence comparison with published adenovirus (AdV) proteins identified the genes for the IIIa, penton, pVII, PVI, 23K proteinase, DNA binding and 100K proteins. The eight EAdV1 genes appeared to be in the same relative order as homologous genes of other AdV. The EAdV1 hexon protein was encoded between the hexon-associated pVI upstream and the 23K proteinase gene downstream and comprised 2742 nucleotides which translated into 913 aa. Similar to other members of the genus Mastadenovirus the EAdV1 hexon yielded two highly conserved genome segments at the N- and C-termini which flanked intermediate variable and hypervariable regions. The majority of the residue differences between EAdV1 and other AdV hexons occurred in two loops that are known for other AdV to protrude from the surface of the nucleocapsid. Amino acid comparisons with other AdV hexons revealed highest homology with HAdV12 hexon with 72% identical and 83% functionally similar residues, followed by bovine AdV3 hexon with 71% identities and 82% functional residue conservation. Phylogenetic analysis suggested that EAdV1 and other AdV do not have an immediate common ancestor.

Phosphorylation of Polyoma and SV40 Virus Proteins

The polypeptides of polyoma and SV40 virions are phosphorylated. An estimate of the amount of phosphorylation of the major virus capsid protein (VPI) has been made using two-dimensional gel electrophoresis to resolve phosphorylated from non-phosphorylated forms. The results suggest that in both polyoma and SV40 virions about 12% of VPI molecules are phosphorylated. In unassembled VPI molecules immunoprecipitated from extracts of infected cells the proportion is greater, about 33%. The possibility that phosphorylated VPI may form the penton proteins of the virus capsid is discussed.

Structure and protein distribution for the capsid of Caulobacter crescentus bacteriophage φCbK

Electron microscope images of negatively stained empty capsids of Caulobacter crescentus bacteriophage φCbK have been analyzed by computer Fourier methods. Two-dimensional computer density maps for the distribution of material in the capsid wall have been obtained by computer Fourier filtering which made possible the complete separation of contributions from the front and back of the capsid including overlapping Fourier coefficients. After scaling for the capsid thickness, using a one-dimensional reconstruction of data from edge-on views of the capsid wall and using topographical information provided by low-angle metal shadowing, a three-dimensional density distribution has been derived. A model for the distribution of subunits in the capsid is proposed which places each of the two major capsid subunit proteins (which are present in a 2:1 ratio) in quasi-equivalent bonding environments. A tentative model is presented in which assembly of the capsid (an elongated T = 7 l icosadeltahedron) is regulated by the bonding geometry of the penton proteins.