Addition of a short peptide ligand to the adenovirus fiber protein.
A major concern associated with the use of recombinant adenoviral vectors is that viral receptors are found on the surface of many cell types and systemic in vivo delivery of the viral vector could result in uncontrolled and widespread expression of therapeutic molecules in many tissues. To construct a cell-type specific recombinant adenoviral vector, a new binding specificity must be added to the virus, and the endogenous binding specificity of the virus must be ablated. In order to introduce a new binding specificity to recombinant adenoviral vectors, the coding sequence of a physiological ligand, the terminal decapeptide of the gastrin releasing peptide (GRP), was placed at the 3' end of the coding sequence of the adenovirus type 5 fiber gene. The resulting fiber-GRP fusion protein was expressed using a T7 vaccinia expression system and has been shown to assemble protein trimers whose quaternary structure is indistinguishable from that of wild-type protein. The fiber-GRP fusion protein was correctly transported to the nucleus of HeLa cells immediately after synthesis. The added GRP ligand in the fiber-GRP fusion protein was accessible to binding by an anti-GRP antibody in both the monomeric and trimeric forms of the chimeric protein. These studies suggest that new cell type specificities for adenovirus binding might be introduced by genetic fusion of peptide ligands on to the carboxyl terminus of the adenovirus fiber protein.
Michael
S I
SI
Gene Therapy Program, University of Alabama at Birmingham 35294-0005, USA.
Hong
J S
JS
Curiel
D T
DT
Engler
J A
JA
D000260:Adenoviruses, Human / Q000235:genetics*; D000595:Amino Acid Sequence; D001483:Base Sequence; D002213:Capsid / Q000737:chemistry / Q000235:genetics*; D036022:Capsid Proteins; D019886:Gastrin-Releasing Peptide; D015870:Gene Expression; D018014:Gene Transfer Techniques; D005814:Genes, Viral / Q000235:genetics; D005822:Genetic Vectors / Q000235:genetics*; D006367:HeLa Cells; D006801:Humans; D008024:Ligands; D008969:Molecular Sequence Data; D010455:Peptides / Q000737:chemistry / Q000235:genetics*; D011487:Protein Conformation; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000737:chemistry; D014162:Transfection; D014616:Vaccinia virus / Q000235:genetics; D015678:Viral Structural Proteins / Q000235:genetics
D036022:Capsid Proteins; D008024:Ligands; D010455:Peptides; D011993:Recombinant Fusion Proteins; D015678:Viral Structural Proteins; C018315:hexon capsid protein, Adenovirus; D019886:Gastrin-Releasing Peptide