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Structural and functional determinants in adenovirus type 2 penton base recombinant protein
1Laboratoire de Virologie et Pathogénèse Moléculaires, Centre National de la Recherche Scientifique URA-1487, Faculté de Médecine, Montpellier, France.
Journal of Virology
|October 29, 1997
Summary
Site-directed mutagenesis revealed key amino acid residues in adenovirus type 2 (Ad2) penton base essential for pentamerization, fiber interaction, and viral functions like cell binding and gene delivery.
Area of Science:
- Virology
- Molecular Biology
- Structural Biology
Background:
- Adenovirus type 2 (Ad2) penton base is crucial for viral structure and function.
- Understanding its discrete domains is key to deciphering viral entry and infection mechanisms.
Purpose of the Study:
- To investigate the roles of specific amino acid residues and domains within the Ad2 penton base.
- To elucidate the structure-function relationships governing penton base-mediated viral processes.
Main Methods:
- Site-directed mutagenesis was employed to generate seventeen substitution mutants of the Ad2 penton base protein.
- Mutant proteins were expressed in baculovirus-infected insect cells and phenotyped in both insect and human cells.
Main Results:
- Penton base pentamerization depends on Trp119, Tyr553, and Lys556.
- Stable fiber-penton base interactions require Arg254, Cys432, Trp439, a basic residue stretch (547-556), and the RGD motif (Arg340).
- Nuclear localization in Sf9 cells was impaired by W119H/W165H mutations and alterations in the polybasic signal (547-549).
- Mutants showed altered HeLa cell binding, cell detachment, DNA internalization, and Ad-mediated gene delivery.
- Integrin-binding motifs (RGD340, LDV287) are functionally linked to other regions (Trp119, Trp165, Cys246, Cys432, Trp439) involved in cell recognition, endocytosis, and post-endocytotic steps.
Conclusions:
- Specific amino acid residues and domains in the Ad2 penton base are critical for its structural integrity and diverse functional roles.
- These findings provide insights into the Ad2 life cycle, from initial cell surface recognition to post-endocytotic events, highlighting the interplay between different domains.