Expression, purification, and characterization of a soluble form of human cytomegalovirus glycoprotein B

Virology
C CarlsonTeresa Compton

Abstract

The human cytomegalovirus glycoprotein B gene (gB; gpUL55) was truncated at amino acid 692 and recombined into Autographa californica nuclear polyhedrosis virus (baculovirus). Infection of insect cells with the recombinant baculovirus resulted in high-level expression and secretion of the truncated gB protein (gB-S) into the culture medium. Purification of gB-S by monoclonal antibody affinity chromatography yielded a protein of ca. 200 kDa. Characterization of the 200-kDa purification product indicated that the recombinant gB protein retained many structural and functional features of the viral gB. Comparison of electrophoretic migration patterns in reduced versus nonreduced protein samples and immune blotting analysis with antibodies specific for the amino or carboxy-terminus of gB demonstrated that the recombinant protein was composed of disulfide linked 69 kDa amino terminal and 35-kDa carboxy-terminal fragments. In addition, recognition of the 200-kDa gB-S by a conformational-dependent, oligomer-specific monoclonal antibody suggested that gB-S was properly folded and dimeric. Like the viral gB, gB-S had heparin binding ability. One heparin binding site was found to reside within the 35-kDa carboxy-terminal fragment (aa 492-...Continue Reading

References

Nov 1, 1969·Archives of Biochemistry and Biophysics·M Steinbuch, R Audran

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Citations

Oct 12, 2005·The Journal of Experimental Medicine·Nagendra R HegdeDavid C Johnson
May 22, 2002·Journal of Virology·Matthew Lopper, Teresa Compton
Oct 21, 2004·Proceedings of the National Academy of Sciences of the United States of America·Adam L FeireTeresa Compton
Dec 3, 2014·Proceedings of the National Academy of Sciences of the United States of America·Anna KabanovaAntonio Lanzavecchia
Mar 24, 2012·Current Opinion in Virology·Adam L Vanarsdall, David C Johnson
Sep 11, 2014·The Journal of Pathology·Paul GriffithsMatt Reeves
Apr 20, 2007·Clinical and Vaccine Immunology : CVI·Elizabeth A ReapJeffrey D Chulay
Dec 14, 2018·Viruses·Christopher C Nguyen, Jeremy P Kamil
Aug 3, 2019·International Journal of Molecular Sciences·Nicolas SmargiassoCatherine Navarre
Sep 24, 2019·Journal of Clinical Virology : the Official Publication of the Pan American Society for Clinical Virology·Paola ZeliniDaniele Lilleri
Jun 22, 2002·Virus Research·Veronika V VrublevskayaOleg S Morenkov

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