title: Active and water-soluble form of lipidated Wnt protein is maintained by a serum glycoprotein afamin/α-albumin.
abstract: Wnt plays important role during development and in various diseases. Because Wnts are lipidated and highly hydrophobic, they can only be purified in the presence of detergents, limiting their use in various in vitro and in vivo assays. We purified N-terminally tagged recombinant Wnt3a secreted from cells and accidentally discovered that Wnt3a co-purified with a glycoprotein afamin derived from the bovine serum included in the media. Wnt3a forms a 1:1 complex with afamin, which remains soluble in aqueous buffer after isolation, and can induce signaling in various cellular systems including the intestical stem cell growth assay. By co-expressing with afamin, biologically active afamin-Wnt complex can be easily obtained in large quantity. As afamin can also solubilize Wnt5a, Wnt3, and many more Wnt subtypes, afamin complexation will open a way to put various Wnt ligands and their signaling mechanisms under a thorough biochemical scrutiny that had been difficult for years.
authors.affiliation: Laboratory of Protein Synthesis and Expression, Institute for Protein Research, Osaka University, Suita, Japan.
authors.forename: Emiko
authors.initials: E
authors.lastname: Mihara
authors.affiliation: Laboratory of Protein Synthesis and Expression, Institute for Protein Research, Osaka University, Suita, Japan.
authors.forename: Hidenori
authors.initials: H
authors.lastname: Hirai
authors.affiliation: Department of Molecular Biology and Biochemistry, Graduate School of Medicine, Osaka University, Suita, Japan.
authors.forename: Hideki
authors.initials: H
authors.lastname: Yamamoto
authors.affiliation: Laboratory of Protein Synthesis and Expression, Institute for Protein Research, Osaka University, Suita, Japan.
authors.forename: Keiko
authors.initials: K
authors.lastname: Tamura-Kawakami
authors.affiliation: Department of Gastroenterology, Keio University School of Medicine, Tokyo, Japan.
authors.forename: Mami
authors.initials: M
authors.lastname: Matano
authors.affiliation: Department of Molecular Biology and Biochemistry, Graduate School of Medicine, Osaka University, Suita, Japan.
authors.forename: Akira
authors.initials: A
authors.lastname: Kikuchi
authors.affiliation: Department of Gastroenterology, Keio University School of Medicine, Tokyo, Japan.
authors.forename: Toshiro
authors.initials: T
authors.lastname: Sato
authors.affiliation: Laboratory of Protein Synthesis and Expression, Institute for Protein Research, Osaka University, Suita, Japan.
authors.forename: Junichi
authors.identifier: http://orcid.org/0000-0002-1219-475X
authors.initials: J
authors.lastname: Takagi
mesh_terms: D000818:Animals; D002352:Carrier Proteins / Q000737:chemistry / Q000378:metabolism*; D002417:Cattle; D006023:Glycoproteins / Q000737:chemistry / Q000378:metabolism*; D057927:Hydrophobic and Hydrophilic Interactions; D011485:Protein Binding; D012709:Serum Albumin / Q000737:chemistry / Q000378:metabolism*; D012995:Solubility; D014867:Water; D051153:Wnt Proteins / Q000737:chemistry / Q000378:metabolism*
chemical_list: D002352:Carrier Proteins; D006023:Glycoproteins; D012709:Serum Albumin; D051153:Wnt Proteins; D014867:Water
keywords: afamin; biochemistry; developmental biology; epitope tag; human; lipid modification; mouse; purification; serum; stem cells; wnt