title: Characterization of the putative native and recombinant rat sterol transporter Niemann-Pick C1 Like 1 (NPC1L1) protein.
abstract: The exact mechanistic pathway of cholesterol absorption in the jejunum of the small intestines is a poorly understood process. Recently, a relatively novel gene, Niemann-Pick C1 Like 1 (NPC1L1), was identified as being critical for intestinal sterol absorption in a pathway which is sensitive to sterol absorption inhibitors such as ezetimibe. NPC1L1 is a multi-transmembrane protein, with a putative sterol sensing domain. Very little else is known about the NPC1L1 protein. In this report, we characterize the native and recombinant rat NPC1L1 protein. We show that NPC1L1 is a 145 kDa membrane protein, enriched in the brush border membrane of the intestinal enterocyte and is highly glycosylated. In addition, sequential detergent extraction of enterocytes result in highly enriched preparations of NPC1L1. An engineered Flag epitope tagged rat NPC1L1 cDNA was expressed as recombinant protein in CHO cells and demonstrated cell surface expression, similar to the native rat protein. These biochemical data indicate that NPC1L1 exists as a predominantly cell surface membrane expressed protein, consistent with its proposed role as the putative intestinal sterol transporter.
authors.affiliation: Department of Cardiovascular/Metabolic Disease, Schering-Plough Research Institute, 2015 Galloping Hill Rd, K15-3-3600, Kenilworth, NJ 07033, USA. sai.iyer@spcorp.com
authors.forename: Sai Prasad N
authors.initials: SP
authors.lastname: Iyer
authors.forename: Xiaorui
authors.initials: X
authors.lastname: Yao
authors.forename: James H
authors.initials: JH
authors.lastname: Crona
authors.forename: Lizbeth M
authors.initials: LM
authors.lastname: Hoos
authors.forename: Glen
authors.initials: G
authors.lastname: Tetzloff
authors.forename: Harry R
authors.initials: HR
authors.lastname: Davis
authors.forename: Michael P
authors.initials: MP
authors.lastname: Graziano
authors.forename: Scott W
authors.initials: SW
authors.lastname: Altmann
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D000911:Antibodies, Monoclonal / Q000276:immunology; D001483:Base Sequence; D017931:DNA Primers; D026901:Membrane Transport Proteins / Q000276:immunology / Q000378:metabolism*; D008969:Molecular Sequence Data; D051381:Rats; D011994:Recombinant Proteins / Q000276:immunology / Q000378:metabolism; D013347:Subcellular Fractions / Q000378:metabolism
chemical_list: D000911:Antibodies, Monoclonal; D017931:DNA Primers; D026901:Membrane Transport Proteins; C482444:NPC1L1 protein, rat; D011994:Recombinant Proteins