Summary auto-generated
This study examined molecular determinants of adenovirus serotype 5 (Ad5) fiber protein binding to its cellular receptor CAR (coxsackie B and adenovirus receptor). Researchers generated multiple mutations in the Ad5 fiber gene, including deletions in the shaft and knob domains and substitutions of Ad5 sequences with Ad3 equivalents. The fiber protein consists of three domains: tail, shaft, and knob, with the knob mediating cell binding. Using baculovirus expression systems, researchers expressed mutant fiber proteins and analyzed their trimerization, assembly with penton base, glycosylation status, and receptor binding capacity. Most monomeric mutants failed to assemble into trimers or form stable penton complexes with the penton base protein, proving lethal for viral replication. However, deletion of strands E and F maintained trimeric structure and assembly capability. Cell binding competition assays using CHO cells expressing CAR revealed that the R-sheet region of the fiber knob contains critical CAR binding motifs. Strands D, G, and I, as well as loops DG and HI, were identified as essential for both trimerization and CAR recognition. Only mutants maintaining proper trimeric structure produced infectious virus particles with reduced but measurable titers. These findings identify specific structural elements required for receptor binding while maintaining viral assembly competence.
Key findings
- Fiber strands D, G, I and loops DG and HI are essential for both trimerization and CAR receptor binding; their deletion prevents virus production
- The R-sheet of the fiber knob monomer contains binding motifs for CAR receptor, with strands E and F regions also involved in receptor recognition
- Fiber monomers fail to assemble with penton base proteins, preventing infectious virion formation, indicating trimerization is prerequisite for assembly
- Only mutations maintaining fiber trimer structure (F5-dl216-314 and F5-dlEF) produced infectious virus with reduced titers, demonstrating assembly requirement for infectivity
- Glycosylation of Ad5 fiber occurs post-trimerization and is not essential for trimerization itself
This summary was generated automatically from the article PDF and is not part of the original publication. Refer to the PDF for the authoritative text.
Abstract
Adenovirus (Ad) tropism is mediated in part through the fibre protein. The common coxsackie B virus and Ad receptor (CAR) was recently identified as the major receptor for subgroup C Ad serotype 5 (Ad5) and serotype 2 (Ad2) fibres. Effects of mutations in the Ad5 fibre gene were studied to assess domains of the fibre capsomer that could alter virus tropism without altering virus assembly and replication. All mutants that accumulated as fibre monomers failed to assemble with a penton base and proved lethal for Ad5 which suggests that the absence of infectious virions resulted in part from a defect in fibre penton base assembly. Cell binding capacity of all fibre mutants was investigated in cell binding competition experiments with adenovirions using CHO--CAR cells (CHO cells that have been transfected with CAR cDNA and express functional CAR). The results suggest that the R-sheet of the Ad5 fibre knob monomer contains binding motifs for CAR and that beta-strands E and F, or a region close to them, may also be involved in receptor recognition.