title: Engineering extended membrane scaffold proteins for self-assembly of soluble nanoscale lipid bilayers.
abstract: High-density lipoproteins (HDLs) play an important role in human health through the metabolism and trafficking of cholesterol as well as providing the feedstocks for steroid hormone biosynthesis. These particles contain proteins, primarily Apo-AI and phospholipid and progress through various structural forms including 'lipid-poor', 'discoidal' and 'spherical' entities as cholesterol esters and lipid are incorporated. The discoidal form of HDL is stabilized in solution by two encircling belts of Apo-AI. Previous protein engineering of the Apo-AI sequence has led to a series of amphipathic helical proteins, termed membrane scaffold proteins (MSPs), which have shown great value in assembling nanoscale soluble membrane bilayers, termed Nanodiscs, of homogeneous size and composition and in the assembly of numerous integral membrane proteins for biophysical and biochemical investigations. In this communication we document a protein engineering approach to generate and optimize an extended polypeptide MSP, which will self-assemble phospholipids into larger Nanodiscs with diameters of 16-17 nm. We extensively characterize these structures by size exclusion chromatography and solution X-ray scattering.
authors.affiliation: School of Molecular and Cellular Biology, University of Illinois, 505 S. Goodwin Avenue, Urbana, IL 61801, USA.
authors.forename: Yelena V
authors.initials: YV
authors.lastname: Grinkova
authors.forename: Ilia G
authors.initials: IG
authors.lastname: Denisov
authors.forename: Stephen G
authors.initials: SG
authors.lastname: Sligar
mesh_terms: D000595:Amino Acid Sequence; D016632:Apolipoprotein A-I / Q000737:chemistry* / Q000235:genetics* / Q000302:isolation & purification; D006801:Humans; D008051:Lipid Bilayers / Q000737:chemistry*; D008075:Lipoproteins, HDL; D008969:Molecular Sequence Data; D049329:Nanostructures / Q000737:chemistry*; D015202:Protein Engineering / Q000379:methods*; D053838:Scattering, Small Angle; D014961:X-Ray Diffraction
chemical_list: D016632:Apolipoprotein A-I; D008051:Lipid Bilayers; D008075:Lipoproteins, HDL
references.citation: Biochemistry. 2007 Feb 27;46(8):2059-69
references.pmid: 17263563
references.citation: Curr Opin Lipidol. 2003 Apr;14(2):151-7
references.pmid: 12642783
references.citation: Proc Natl Acad Sci U S A. 2002 May 14;99(10):6725-30
references.pmid: 11997441
references.citation: J Phys Chem B. 2007 Sep 27;111(38):11095-104
references.pmid: 17696388
references.citation: Nano Lett. 2007 Jun;7(6):1692-6
references.pmid: 17503871
references.citation: Biochem Biophys Res Commun. 2003 Dec 5;312(1):115-9
references.pmid: 14630028
references.citation: Protein Sci. 2003 Nov;12(11):2476-81
references.pmid: 14573860
references.citation: Biochemistry. 1988 Oct 18;27(21):8261-9
references.pmid: 3233209
references.citation: J Struct Biol. 1998 Sep;123(1):37-44
references.pmid: 9774543
references.citation: J Mol Med (Berl). 2006 Apr;84(4):276-94
references.pmid: 16501936
references.citation: Protein Sci. 1995 Nov;4(11):2411-23
references.pmid: 8563639
references.citation: FEBS Lett. 2010 May 3;584(9):1721-7
references.pmid: 19836392
references.citation: Biotechniques. 2003 Sep;35(3):556-60, 562-3
references.pmid: 14513561
references.citation: Biochim Biophys Acta. 2011 Jan;1814(1):223-9
references.pmid: 20685623
references.citation: J Am Chem Soc. 2004 Mar 24;126(11):3477-87
references.pmid: 15025475
references.citation: J Phys Chem B. 2005 Aug 18;109(32):15580-8
references.pmid: 16852976
references.citation: Cell Mol Life Sci. 2001 Jun;58(7):885-93
references.pmid: 11497237