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Detergent extraction of herpes simplex virus type 1 glycoprotein D by zwitterionic and non-ionic detergents and purification by ion-exchange high-performance liquid chromatography

1998; Elsevier BV; Volume: 816; Issue: 1 Linguagem: Inglês

10.1016/s0021-9673(98)00288-x

1873-3778

Sytske Welling‐Wester, Matty Feijlbrief, Danny G. A. M. Koedijk, Gjalt W. Welling,

Toxin Mechanisms and Immunotoxins

Detergents (surfactants) are the key reagents in the extraction and purification of integral membrane proteins. Zwitterionic and non-ionic detergents were used for the extraction of recombinant glycoprotein D (gD-1) of herpes simplex virus type 1 (HSV-1) from insect cells infected with recombinant baculovirus. The highest yield was obtained with the two alkyl carboxybetaine detergents (N-dodecyl-N,N-dimethylammonio)undecanoate [DDMAU, critical micelle concentration (CMC)=0.13 mM] and (N-dodecyl-N,N-dimethylammonio)butyrate (DDMAB, CMC=4.3 mM). Therefore these zwitterionic detergents were used as additives to the elution buffers in ion-exchange high-performance liquid chromatography (HPIEC) to purify gD-1 of HSV-1 from the extracts. The non-ionic detergent pentaethyleneglycol monodecyl ether (C10E5) that was used in earlier studies [R.A. Damhof, M. Feijlbrief, S. Welling-Wester, G.W. Welling, J. Chromatogr. A, 676 (1994) 43] was used for comparison. Two columns were used, Mono Q and Resource Q, at 1 and 5 ml/min flow-rates, respectively. The results show that the detergents DDMAU and C10E5 are superior to DDMAB, when the detergents were used as additives to the elution buffers at 0.2% (w/v). With 0.2% DDMAB in the eluent, purification of HSV gD-1 was not possible. Detergents with a high CMC may be less suitable as additives in elution buffers. HPIEC at flow-rates of 1 and at 5 ml/min showed satisfactory results. At 5 ml/min HSV gD-1 was mainly concentrated in two eluent fractions. The highest recovery of gD-1 was obtained either by chromatography of a C10E5 extract using a Mono Q column at a flow-rate of 1 ml/min or by chromatography of a DDMAU extract using a Resource Q column at a flow-rate of 5 ml/min.