title: Structure and stability of the spinach aquaporin SoPIP2;1 in detergent micelles and lipid membranes.
abstract: BACKGROUND
  SoPIP2;1 constitutes one of the major integral proteins in spinach leaf plasma membranes and belongs to the aquaporin family. SoPIP2;1 is a highly permeable and selective water channel that has been successfully overexpressed and purified with high yields. In order to optimize reconstitution of the purified protein into biomimetic systems, we have here for the first time characterized the structural stability of SoPIP2;1.
  
  
  RESULTS
  We have characterized the protein structural stability after purification and after reconstitution into detergent micelles and proteoliposomes using circular dichroism and fluorescence spectroscopy techniques. The structure of SoPIP2;1 was analyzed either with the protein solubilized with octyl-β-D-glucopyranoside (OG) or reconstituted into lipid membranes formed by E. coli lipids, diphytanoylphosphatidylcholine (DPhPC), or reconstituted into lipid membranes formed from mixtures of 1-palmitoyl-2-oleoyl-phosphatidylcholine (POPE), 1-palmitoyl-2oleoyl-phosphatidylethanolamine (POPE), 1-palmitoyl-2-oleoyl-phosphatidylserine (POPS), and ergosterol. Generally, SoPIP2;1 secondary structure was found to be predominantly α-helical in accordance with crystallographic data. The protein has a high thermal structural stability in detergent solutions, with an irreversible thermal unfolding occurring at a melting temperature of 58°C. Incorporation of the protein into lipid membranes increases the structural stability as evidenced by an increased melting temperature of up to 70°C.
  
  
  CONCLUSIONS
  The results of this study provide insights into SoPIP2;1 stability in various host membranes and suggest suitable choices of detergent and lipid composition for reconstitution of SoPIP2;1 into biomimetic membranes for biotechnological applications.
authors.affiliation: Department of Physics and Chemistry, MEMPHYS-Center for Biomembrane Physics, University of Southern Denmark, Odense, Denmark. miplsen@memphys.sdu.dk
authors.forename: Inés
authors.initials: I
authors.lastname: Plasencia
authors.forename: Sabeen
authors.initials: S
authors.lastname: Survery
authors.forename: Sania
authors.initials: S
authors.lastname: Ibragimova
authors.forename: Jesper S
authors.initials: JS
authors.lastname: Hansen
authors.forename: Per
authors.initials: P
authors.lastname: Kjellbom
authors.forename: Claus
authors.initials: C
authors.lastname: Helix-Nielsen
authors.forename: Urban
authors.initials: U
authors.lastname: Johanson
authors.forename: Ole G
authors.initials: OG
authors.lastname: Mouritsen
mesh_terms: D020346:Aquaporins / Q000032:analysis / Q000737:chemistry* / Q000187:drug effects / Q000378:metabolism*; D002462:Cell Membrane / Q000502:physiology*; D002942:Circular Dichroism; D003902:Detergents / Q000494:pharmacology*; D008563:Membrane Lipids / Q000494:pharmacology*; D008823:Micelles; D011489:Protein Denaturation / Q000187:drug effects; D055550:Protein Stability / Q000187:drug effects; D017433:Protein Structure, Secondary / Q000187:drug effects; D017434:Protein Structure, Tertiary / Q000187:drug effects; D058767:Protein Unfolding / Q000187:drug effects; D011510:Proteolipids / Q000494:pharmacology / Q000502:physiology; D018724:Spinacia oleracea / Q000737:chemistry* / Q000378:metabolism; D013329:Structure-Activity Relationship; D013696:Temperature
chemical_list: D020346:Aquaporins; D003902:Detergents; D008563:Membrane Lipids; D008823:Micelles; D011510:Proteolipids; C020485:proteoliposomes