@article {Qi:2009:0960-7412:932,
title = "Purification of low-abundance Arabidopsis plasma-membrane protein complexes and identification of candidate components",
journal = "The Plant Journal",
parent_itemid = "infobike://bsc/tpj",
publishercode ="bp",
year = "2009",
volume = "57",
number = "5",
publication date ="2009-03-01T00:00:00",
pages = "932-944",
itemtype = "ARTICLE",
issn = "0960-7412",
eissn = "1365-313X",
url = "https://www.ingentaconnect.com/content/bsc/tpj/2009/00000057/00000005/art00014",
doi = "doi:10.1111/j.1365-313X.2008.03736.x",
keyword = "plasma membrane, HPB tag, RPS2, biotinylation, protein complexes",
author = "Qi, Yiping and Katagiri, Fumiaki",
abstract = "Summary Purification of low-abundance plasma-membrane (PM) protein complexes is a challenging task. We devised a tandem affinity purification tag termed the HPB tag, which contains the biotin carboxyl carrier protein domain (BCCD) of Arabidopsis 3-methylcrotonalCoA carboxylase. The BCCD is biotinylated in vivo, and the tagged protein can be captured by streptavidin beads. All five C-terminally tagged Arabidopsis proteins tested, including four PM proteins, were functional and biotinylated with high efficiency in Arabidopsis. Transgenic Arabidopsis plants expressing an HPB-tagged protein, RPS2::HPB, were used to develop a method to purify protein complexes containing the HPB-tagged protein. RPS2 is a membrane-associated disease resistance protein of low abundance. The purification method involves microsomal fractionation, chemical cross-linking, solubilization, and one-step affinity purification using magnetic streptavidin beads, followed by protein identification using LC-MS/MS. We identified RIN4, a known RPS2 interactor, as well as other potential components of the RPS2 complex(es). Thus, the HPB tag method is suitable for the purification of low-abundance PM protein complexes.",
}