title: An efficient tag derived from the common epitope of tospoviral NSs proteins for monitoring recombinant proteins expressed in both bacterial and plant systems.
abstract: NSscon (23 aa), a common epitope in the gene silencing suppressor NSs proteins of the members of the Watermelon silver mottle virus (WSMoV) serogroup, was previously identified. In this investigation, we expressed different green fluorescent protein (GFP)-fused deletions of NSscon in bacteria and reacted with NSscon monoclonal antibody (MAb). Our results indicated that the core 9 amino acids, "(109)KFTMHNQIF(117)", denoted as "nss", retain the reactivity of NSscon. In bacterial pET system, four different recombinant proteins labeled with nss, either at N- or C-extremes, were readily detectable without position effects, with sensitivity superior to that for the polyhistidine-tag. When the nss-tagged Zucchini yellow mosaic virus (ZYMV) helper component-protease (HC-Pro) and WSMoV nucleocapsid protein were transiently expressed by agroinfiltration in tobacco, they were readily detectable and the tag's possible efficacy for gene silencing suppression was not noticed. Co-immunoprecipitation of nss-tagged and non-tagged proteins expressed from bacteria confirmed the interaction of potyviral HC-Pro and coat protein. Thus, we conclude that this novel nss sequence is highly valuable for tagging recombinant proteins in both bacterial and plant expression systems.
authors: Department of Plant Pathology, National Chung Hsing University, Taichung 40227, Taiwan, ROC.
authors: Hao-Wen
authors: HW
authors: Cheng
authors: Kuan-Chun
authors: KC
authors: Chen
authors: Joseph A J
authors: JA
authors: Raja
authors: Jian-Xian
authors: JX
authors: Li
authors: Shyi-Dong
authors: SD
authors: Yeh
mesh_terms: D000595:Amino Acid Sequence; D001709:Biotechnology; D015153:Blotting, Western; D028464:Cucurbita / Q000235:genetics; D000939:Epitopes / Q000737:chemistry / Q000276:immunology / Q000378:metabolism*; D004926:Escherichia coli / Q000235:genetics; D049452:Green Fluorescent Proteins; D047468:Immunoprecipitation; D008969:Molecular Sequence Data; D009029:Mosaic Viruses / Q000235:genetics; D009842:Oligopeptides / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D018515:Plant Leaves / Q000737:chemistry / Q000378:metabolism; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification* / Q000378:metabolism*; D017865:Tospovirus / Q000235:genetics*; D017361:Viral Nonstructural Proteins / Q000737:chemistry / Q000235:genetics / Q000276:immunology / Q000378:metabolism*
chemical_list: D000939:Epitopes; D009842:Oligopeptides; D011993:Recombinant Fusion Proteins; D017361:Viral Nonstructural Proteins; D049452:Green Fluorescent Proteins