Purification of the integral membrane glycoproteins D of herpes simplex virus types 1 and 2, produced in the recombinant baculovirus expression system, by ion-exchange high-performance liquid chromatography.
Selective elution of Sendai virus integral membrane proteins by ion-exchange high-performance liquid chromatography (HPIEC) using different detergent concentrations was reported before [S. Welling-Wester, M. Freijlbrief, D.G.A.M. Koedijk, M.A. Braaksma, B.R.K. Douma and G.W. Welling, J. Chromatogr., 646 (1993) 37]. In the present study this novel approach was applied to the purification of the integral membrane glycoprotein D of Herpes simplex virus type 1 and 2. The glycoproteins D of types 1 (gD-1) and 2 (gD-2) were cloned into the baculovirus expression system and produced in protein-free cultured insect cells. Detergent extracts of recombinant baculovirus-infected insect cells containing gD-1 or gD-2 were prepared using pentaethyleneglycol monodecyl ether, for extraction (final concentration 2%, w/v). The same detergent was used as additive in the elution buffers for HPIEC on a Mono Q HR 5/5 column. At low (0.005%) detergent concentration, most of the proteins present in the extract including part of gD were eluted with the sodium chloride gradient whereas a subsequent blank run using the same gradient at higher detergent concentration (0.1%) resulted in selective elution of pure gD.
Damhof
R A
RA
Laboratorium voor Medische Microbiologie, Rijksuniversiteit Groningen, Netherlands.
Feijlbrief
M
M
Welling-Wester
S
S
Welling
G W
GW
D000818:Animals; D016367:Baculoviridae / Q000235:genetics / Q000378:metabolism*; D002851:Chromatography, High Pressure Liquid / Q000379:methods*; D002852:Chromatography, Ion Exchange; D003902:Detergents; D004591:Electrophoresis, Polyacrylamide Gel; D004797:Enzyme-Linked Immunosorbent Assay; D018259:Herpesvirus 1, Human / Q000737:chemistry*; D018258:Herpesvirus 2, Human / Q000737:chemistry*; D009036:Moths; D011994:Recombinant Proteins / Q000302:isolation & purification; D014759:Viral Envelope Proteins / Q000302:isolation & purification*
D003902:Detergents; D011994:Recombinant Proteins; D014759:Viral Envelope Proteins; C039447:glycoprotein D, Human herpesvirus 1; C040570:glycoprotein D-herpes simplex virus type 2