Expression, glycosylation, and modification of the spike (S) glycoprotein of SARS CoV.
The spike (S) glycoprotein of coronaviruses is known to be essential in the binding of the virus to the host cell at the advent of the infection process. To study the maturation pathway of the S glycoprotein of the severe acute respiratory syndrome (SARS)-coronavirus (CoV) within the host cell, a T7/vaccinia virus-based expression system coupled to immunoprecipitation with anti-S antibodies was used to test and analyze different forms of the S glycoprotein. The state of maturity of the S glycoprotein can be deduced from its sensitivity to hydrolysis by endoglycosidase H (EndoH) or N-glycosidase F (N-Gly F). A fully matured S glycoprotein will be modified with complex oligosaccharides which makes it resistant to cleavage by EndoH but not by N-Gly F. By exploiting this characteristic, it is then possible to determine which forms of the immunoprecipitated S protein are properly processed by the host cell. With this system, many different constructs of the S glycoprotein can be analyzed in parallel thus providing another method by which to study the functional domains of S involved in membrane fusion event that occurs during viral infection.
Shen
Shuo
S
Collaborative Antiviral Research Group, Institute of Molecular and Cell Biology, Proteos, Singapore.
Tan
Timothy H P
TH
Tan
Yee-Joo
YJ
D000818:Animals; D019556:COS Cells; D002522:Chlorocebus aethiops; D015870:Gene Expression; D006031:Glycosylation; D017038:Mannosyl-Glycoprotein Endo-beta-N-Acetylglucosaminidase / Q000737:chemistry; D008562:Membrane Glycoproteins / Q000032:analysis / Q000096:biosynthesis* / Q000235:genetics; D043524:Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase / Q000737:chemistry; D046188:Protein Modification, Translational / Q000502:physiology*; D011994:Recombinant Proteins / Q000032:analysis / Q000096:biosynthesis* / Q000235:genetics; D045473:Severe acute respiratory syndrome-related coronavirus / Q000235:genetics; D064370:Spike Glycoprotein, Coronavirus; D014616:Vaccinia virus / Q000235:genetics; D014709:Vero Cells; D014759:Viral Envelope Proteins / Q000032:analysis / Q000096:biosynthesis* / Q000235:genetics
D008562:Membrane Glycoproteins; D011994:Recombinant Proteins; D064370:Spike Glycoprotein, Coronavirus; D014759:Viral Envelope Proteins; C578557:spike glycoprotein, SARS-CoV; C578553:spike protein, mouse hepatitis virus; D017038:Mannosyl-Glycoprotein Endo-beta-N-Acetylglucosaminidase; D043524:Peptide-N4-(N-acetyl-beta-glucosaminyl) Asparagine Amidase