Functional characterization of the Sindbis virus E2 glycoprotein by transposon linker-insertion mutagenesis.
The glycoprotein envelope of alphaviruses consists of two proteins, E1 and E2. E1 is responsible for fusion and E2 is responsible for receptor binding. An atomic structure is available for E1, but one for E2 has not been reported. In this study, transposon linker-insertion mutagenesis was used to probe the function of different domains of E2. A library of mutants, containing 19 amino acid insertions in the E2 glycoprotein sequence of the prototype alphavirus, Sindbis virus (SINV), was generated. Fifty-seven independent E2 insertions were characterized, of which more than half (67%) gave rise to viable virus. The wild-type-like mutants identify regions that accommodate insertions without perturbing virus production and can be used to insert targeting moieties to direct SINV to specific receptors. The defective and lethal mutants give insight into regions of E2 important for protein stability, transport to the cell membrane, E1-E2 contacts, and receptor binding.
Navaratnarajah
Chanakha K
CK
Department of Biological Sciences, Purdue University, 915 W. State Street, West Lafayette, IN 47907-2054, USA.
Kuhn
Richard J
RJ
D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D006224:Cricetinae; D004251:DNA Transposable Elements / Q000235:genetics*; D008969:Molecular Sequence Data; D016254:Mutagenesis, Insertional / Q000235:genetics*; D009500:Neutralization Tests; D016366:Open Reading Frames / Q000235:genetics; D021381:Protein Transport; D012845:Sindbis Virus / Q000235:genetics* / Q000472:pathogenicity / Q000502:physiology; D014759:Viral Envelope Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*
D000911:Antibodies, Monoclonal; D004251:DNA Transposable Elements; D014759:Viral Envelope Proteins; C064950:glycoprotein E2, Sindbis virus