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Size-exclusion chromatographic study of the reduction of recombinant hepatitis B surface antigen

机译:大小排阻色谱法研究重组乙型肝炎表面抗原的减少

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The reduction of the P. pastoris-derived hepatitis B surface antigen (HBsAg) has been investigated by size exclusion chromatography performed in a detergent solution containing 0.3% sodium dodecyl sulfate (SDS) and 0.1 M Tris-HCl, pH 7.0. The HBsAg, reduced under different conditions and passed through the TSK G4000 SW column (600x7.5 mm I.D.) at 0.9 ml min(-1), was resolved into two peaks corresponding to the reduced, monomeric, and non-reduced forms, respectively. Under these conditions, the antigen fraction corresponding to the HBsAg dimer can be separated and completely reduced to monomers by repeated reductive treatment with simultaneous lipid removal. The efficiency of reduction was maximal after sample treatment with an equal volume of a solution containing 417 mM dithiothreitol, 4.2% (w/v) SDS and 16% (v/v) 2-mercaptoethanol. In conclusion, complete reduction of recombinant HBsAg to monomer subunits is possible and depends on the efficiency of lipid removal during the reductive treatment. (C) 1998 Elsevier Science B.V. All rights reserved. [References: 19]
机译:已经通过在含有0.3%十二烷基硫酸钠(SDS)和0.1 M Tris-HCl,pH 7.0的去污剂溶液中进行的尺寸排阻色谱法研究了巴斯德毕赤酵母来源的乙型肝炎表面抗原(HBsAg)的减少。在不同条件下还原并以0.9 ml min(-1)通过TSK G4000 SW色谱柱(600x7.5 mm内径)的HBsAg被拆分为两个峰,分别对应于还原形式,单体形式和非还原形式。在这些条件下,可以通过重复还原处理并同时去除脂质,分离出与HBsAg二聚体相对应的抗原部分并将其完全还原为单体。用等体积的含417 mM二硫苏糖醇,4.2%(w / v)SDS和16%(v / v)2-巯基乙醇的溶液处理样品后,还原效率最高。总之,重组HBsAg完全还原成单体亚基是可能的,并且取决于还原处理过程中脂质去除的效率。 (C)1998 Elsevier Science B.V.保留所有权利。 [参考:19]

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