Skip to main content
NCBI home page
Protein Science : A Publication of the Protein Society logoLink to Protein Science : A Publication of the Protein Society
. 2000 Dec;9(12):2518–2527. doi: 10.1110/ps.9.12.2518

An improved tripod amphiphile for membrane protein solubilization.

S M Yu 1, D T McQuade 1, M A Quinn 1, C P Hackenberger 1, M P Krebs 1, A S Polans 1, S H Gellman 1
PMCID: PMC2144526  PMID: 11206073

Abstract

Intrinsic membrane proteins represent a large fraction of the proteins produced by living organisms and perform many crucial functions. Structural and functional characterization of membrane proteins generally requires that they be extracted from the native lipid bilayer and solubilized with a small synthetic amphiphile, for example, a detergent. We describe the development of a small molecule with a distinctive amphiphilic architecture, a "tripod amphiphile," that solubilizes both bacteriorhodopsin (BR) and bovine rhodopsin (Rho). The polar portion of this amphiphile contains an amide and an amine-oxide; small variations in this polar segment are found to have profound effects on protein solubilization properties. The optimal tripod amphiphile extracts both BR and Rho from the native membrane environments and maintains each protein in a monomeric native-like form for several weeks after delipidation. Tripod amphiphiles are designed to display greater conformational rigidity than conventional detergents, with the long-range goal of promoting membrane protein crystallization. The results reported here represent an important step toward that ultimate goal.

Full Text

The Full Text of this article is available as a PDF (513.2 KB).

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Bakker E. P., Caplan S. R. Phospholipid substitution of the purple membrane. The stoichiometry of light-induced proton release by phospholipid-substituted purple membranes. Biochim Biophys Acta. 1978 Aug 8;503(2):362–379. doi: 10.1016/0005-2728(78)90194-9. [DOI] [PubMed] [Google Scholar]
  2. Chattopadhyay A., London E. Fluorimetric determination of critical micelle concentration avoiding interference from detergent charge. Anal Biochem. 1984 Jun;139(2):408–412. doi: 10.1016/0003-2697(84)90026-5. [DOI] [PubMed] [Google Scholar]
  3. Corcelli A., Lobasso S., Colella M., Trotta M., Guerrieri A., Palmisano F. Role of palmitic acid on the isolation and properties of halorhodopsin. Biochim Biophys Acta. 1996 Jun 11;1281(2):173–181. doi: 10.1016/0005-2736(96)00007-7. [DOI] [PubMed] [Google Scholar]
  4. De Grip W. J. Purification of bovine rhodopsin over concanavalin A--sepharose. Methods Enzymol. 1982;81:197–207. doi: 10.1016/s0076-6879(82)81032-x. [DOI] [PubMed] [Google Scholar]
  5. De Grip W. J. Thermal stability of rhodopsin and opsin in some novel detergents. Methods Enzymol. 1982;81:256–265. doi: 10.1016/s0076-6879(82)81040-9. [DOI] [PubMed] [Google Scholar]
  6. Dencher N. A., Heyn M. P. Formation and properties of bacteriorhodopsin monomers in the non-ionic detergents octyl-beta-D-glucoside and Triton X-100. FEBS Lett. 1978 Dec 15;96(2):322–326. doi: 10.1016/0014-5793(78)80427-x. [DOI] [PubMed] [Google Scholar]
  7. Garavito R. M., Picot D., Loll P. J. Strategies for crystallizing membrane proteins. J Bioenerg Biomembr. 1996 Feb;28(1):13–27. [PubMed] [Google Scholar]
  8. Hansen J. C., Lebowitz J., Demeler B. Analytical ultracentrifugation of complex macromolecular systems. Biochemistry. 1994 Nov 15;33(45):13155–13163. doi: 10.1021/bi00249a001. [DOI] [PubMed] [Google Scholar]
  9. Heyn M. P., Bauer P. J., Dencher N. A. A natural CD label to probe the structure of the purple membrane from Halobacterium halobium by means of exciton coupling effects. Biochem Biophys Res Commun. 1975 Dec 1;67(3):897–903. doi: 10.1016/0006-291x(75)90761-5. [DOI] [PubMed] [Google Scholar]
  10. Hjelmeland L. M., Nebert D. W., Osborne J. C., Jr Sulfobetaine derivatives of bile acids: nondenaturing surfactants for membrane biochemistry. Anal Biochem. 1983 Apr 1;130(1):72–82. doi: 10.1016/0003-2697(83)90651-6. [DOI] [PubMed] [Google Scholar]
  11. Khorana H. G. Two light-transducing membrane proteins: bacteriorhodopsin and the mammalian rhodopsin. Proc Natl Acad Sci U S A. 1993 Feb 15;90(4):1166–1171. doi: 10.1073/pnas.90.4.1166. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Kühlbrandt W. Three-dimensional crystallization of membrane proteins. Q Rev Biophys. 1988 Nov;21(4):429–477. doi: 10.1017/s0033583500004625. [DOI] [PubMed] [Google Scholar]
  13. Lau F. W., Bowie J. U. A method for assessing the stability of a membrane protein. Biochemistry. 1997 May 13;36(19):5884–5892. doi: 10.1021/bi963095j. [DOI] [PubMed] [Google Scholar]
  14. Luecke H., Schobert B., Richter H. T., Cartailler J. P., Lanyi J. K. Structure of bacteriorhodopsin at 1.55 A resolution. J Mol Biol. 1999 Aug 27;291(4):899–911. doi: 10.1006/jmbi.1999.3027. [DOI] [PubMed] [Google Scholar]
  15. McClare C. W. An accurate and convenient organic phosphorus assay. Anal Biochem. 1971 Feb;39(2):527–530. doi: 10.1016/0003-2697(71)90443-x. [DOI] [PubMed] [Google Scholar]
  16. McQuade DT, Quinn MA, Yu SM, Polans AS, Krebs MP, Gellman SH. Rigid Amphiphiles for Membrane Protein Manipulation. Angew Chem Int Ed Engl. 2000 Feb;39(4):758–761. [PubMed] [Google Scholar]
  17. Miercke L. J., Ross P. E., Stroud R. M., Dratz E. A. Purification of bacteriorhodopsin and characterization of mature and partially processed forms. J Biol Chem. 1989 May 5;264(13):7531–7535. [PubMed] [Google Scholar]
  18. Møller J. V., le Maire M. Detergent binding as a measure of hydrophobic surface area of integral membrane proteins. J Biol Chem. 1993 Sep 5;268(25):18659–18672. [PubMed] [Google Scholar]
  19. Oesterhelt D., Stoeckenius W. Isolation of the cell membrane of Halobacterium halobium and its fractionation into red and purple membrane. Methods Enzymol. 1974;31:667–678. doi: 10.1016/0076-6879(74)31072-5. [DOI] [PubMed] [Google Scholar]
  20. Ostermeier C., Michel H. Crystallization of membrane proteins. Curr Opin Struct Biol. 1997 Oct;7(5):697–701. doi: 10.1016/s0959-440x(97)80080-2. [DOI] [PubMed] [Google Scholar]
  21. Palczewski K., Kumasaka T., Hori T., Behnke C. A., Motoshima H., Fox B. A., Le Trong I., Teller D. C., Okada T., Stenkamp R. E. Crystal structure of rhodopsin: A G protein-coupled receptor. Science. 2000 Aug 4;289(5480):739–745. doi: 10.1126/science.289.5480.739. [DOI] [PubMed] [Google Scholar]
  22. Papermaster D. S. Preparation of retinal rod outer segments. Methods Enzymol. 1982;81:48–52. doi: 10.1016/s0076-6879(82)81010-0. [DOI] [PubMed] [Google Scholar]
  23. Reyenolds J. A., Stoeckenius W. Molecular weight of bacteriorhodopsin solubilized in Triton X-100. Proc Natl Acad Sci U S A. 1977 Jul;74(7):2803–2804. doi: 10.1073/pnas.74.7.2803. [DOI] [PMC free article] [PubMed] [Google Scholar]
  24. Sakmar T. P. Rhodopsin: a prototypical G protein-coupled receptor. Prog Nucleic Acid Res Mol Biol. 1998;59:1–34. doi: 10.1016/s0079-6603(08)61027-2. [DOI] [PubMed] [Google Scholar]
  25. Schafmeister C. E., Miercke L. J., Stroud R. M. Structure at 2.5 A of a designed peptide that maintains solubility of membrane proteins. Science. 1993 Oct 29;262(5134):734–738. doi: 10.1126/science.8235592. [DOI] [PubMed] [Google Scholar]
  26. Schertler G. F., Bartunik H. D., Michel H., Oesterhelt D. Orthorhombic crystal form of bacteriorhodopsin nucleated on benzamidine diffracting to 3.6 A resolution. J Mol Biol. 1993 Nov 5;234(1):156–164. doi: 10.1006/jmbi.1993.1570. [DOI] [PubMed] [Google Scholar]
  27. Tanford C., Reynolds J. A. Characterization of membrane proteins in detergent solutions. Biochim Biophys Acta. 1976 Oct 26;457(2):133–170. doi: 10.1016/0304-4157(76)90009-5. [DOI] [PubMed] [Google Scholar]
  28. Timmins P., Pebay-Peyroula E., Welte W. Detergent organisation in solutions and in crystals of membrane proteins. Biophys Chem. 1994 Dec;53(1-2):27–36. doi: 10.1016/0301-4622(94)00073-5. [DOI] [PubMed] [Google Scholar]
  29. Tribet C., Audebert R., Popot J. L. Amphipols: polymers that keep membrane proteins soluble in aqueous solutions. Proc Natl Acad Sci U S A. 1996 Dec 24;93(26):15047–15050. doi: 10.1073/pnas.93.26.15047. [DOI] [PMC free article] [PubMed] [Google Scholar]
  30. Wallin E., von Heijne G. Genome-wide analysis of integral membrane proteins from eubacterial, archaean, and eukaryotic organisms. Protein Sci. 1998 Apr;7(4):1029–1038. doi: 10.1002/pro.5560070420. [DOI] [PMC free article] [PubMed] [Google Scholar]
  31. White S. H., Wimley W. C. Membrane protein folding and stability: physical principles. Annu Rev Biophys Biomol Struct. 1999;28:319–365. doi: 10.1146/annurev.biophys.28.1.319. [DOI] [PubMed] [Google Scholar]
  32. Zhou Y., Bowie J. U. Building a thermostable membrane protein. J Biol Chem. 2000 Mar 10;275(10):6975–6979. doi: 10.1074/jbc.275.10.6975. [DOI] [PubMed] [Google Scholar]

Articles from Protein Science : A Publication of the Protein Society are provided here courtesy of The Protein Society

RESOURCES