A simple procedure for expression and purification of selected non-structural (alpha and beta) herpes simplex virus 1 (HSV-1) proteins.
The expression and isolation of herpes simplex virus 1 (HSV-1) immediate early (alpha) IE63 (ICP27) and of the early (beta) thymidine kinase (Tk) polypeptides in Escherichia coli JM 109 cells transformed with the PinPoint Xa-1 (Promega) plasmid construct carrying either the HSV-1 UL54 or UL23 genes are described. The resulting biotinylated fusion protein(s) could be easily induced and were purified in appropriate amounts by means of a monomeric avidin-conjugated resin (SoftLink Soft Release Avidin Resin, Promega) provided that: (1) the exponential growth of the selected transformed cells was monitored carefully; (2) the post-induction harvest interval was properly chosen; and (3) the period for adsorption to the avidin resin suitably adjusted. The isolated protein(s), although partially digested in the case of the IE63 polypeptide, were suitable antigen(s) for immunization of various animal species. Co-purification of trace amounts of endogenous biotinylated protein(s) produced in E. coli was eliminated by shortening the duration of adsorption to the avidin resin.
Institute of Virology, Slovak Academy of Sciences, Dúbravská cesta 9, 842 45 4, Bratislava, Slovak Republic.
J
J
Kosovský
V
V
Durmanová
M
M
Kúdelová
I
I
Rezuchová
L
L
Tkáciková
J
J
Rajcáni
D004591:Electrophoresis, Polyacrylamide Gel; D004926:Escherichia coli / Q000235:genetics / Q000254:growth & development / Q000378:metabolism*; D018259:Herpesvirus 1, Human / Q000235:genetics* / Q000378:metabolism; D017874:Immediate-Early Proteins / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism*; D010957:Plasmids / Q000235:genetics; D011993:Recombinant Fusion Proteins / Q000378:metabolism; D013937:Thymidine Kinase / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism*; D014170:Transformation, Genetic
C057120:ICP27 protein, human herpesvirus 1; D017874:Immediate-Early Proteins; D011993:Recombinant Fusion Proteins; D013937:Thymidine Kinase; C092717:thymidine kinase 1