Purification by Strep-Tactin affinity chromatography of a delete envelope gp51 protein of Bovine Leukaemia virus expressed in Sf21 insect cells.
Bovine leukaemia virus (BLV) causes disease in cattle and it is related to human T lymphotrofic viruses HTLV-1 and HTLV-2. The objective of this study was to express and purify deleted and stable forms of the gp51 envelope glycoprotein of BLV using a baculovirus system. Two forms of the gp51 were synthesised: one comprised the gp51 N-terminal 174 amino acids and a single 6xHis tag (Delta(175-268)gp51-His) and the second form contained the same amino acid sequence and a C-terminal Strep-tag II in addition to the 6xHis tag (Delta(175-268)gp51-STH). The two proteins were expressed and purified by immobilized metal-affinity chromatography (IMAC) or by Strep-Tactin column. The Strep-Tactin technology was more efficient than IMAC method and achieved a high pure recombinant deleted gp51. In addition the Delta(175-268)gp51-STH protein was further concentrated by IMAC. This purified antigen could be used for the isolation of immunoreactive molecules and to develop a competitive ELISA test.
De Giuseppe
Antonio
A
Istituto Zooprofilattico Sperimentale dell'Umbria e delle Marche, G. Salvemini 1, 06126, Perugia, Italy. a.degiuseppe@izsum.it
Forti
Katia
K
Feliziani
Francesco
F
Severi
Giulio
G
Cagiola
Monica
M
D000818:Animals; D016367:Baculoviridae / Q000235:genetics; D015153:Blotting, Western; D002846:Chromatography, Affinity / Q000379:methods*; D004591:Electrophoresis, Polyacrylamide Gel; D006639:Histidine / Q000235:genetics / Q000378:metabolism; D001909:Leukemia Virus, Bovine / Q000737:chemistry* / Q000235:genetics; D009842:Oligopeptides / Q000235:genetics / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000096:biosynthesis / Q000235:genetics / Q000302:isolation & purification*; D018411:Spodoptera / Q000378:metabolism*; D014759:Viral Envelope Proteins / Q000096:biosynthesis / Q000235:genetics / Q000302:isolation & purification*
C099517:Ala-Trp-Arg-His-Pro-Gln-Phe-Gly-Gly; C471213:His-His-His-His-His-His; D009842:Oligopeptides; D011993:Recombinant Fusion Proteins; D014759:Viral Envelope Proteins; D006639:Histidine