How to make tubular crystals by reconstitution of detergent-solubilized Ca2(+)-ATPase

Biophysical Journal
H S YoungDavid L Stokes

Abstract

In an attempt to better define the parameters governing reconstitution and two-dimensional crystallization of membrane proteins, we have studied Ca2(+)-ATPase from rabbit sarcoplasmic reticulum. This ion pump forms vanadate-induced crystals in its native membrane and has previously been reconstituted at high lipid-to-protein ratios for functional studies. We have characterized the reconstitution of purified Ca2(+)-ATPase at low lipid-to-protein ratios and discovered procedures that produce long, tubular crystals suitable for helical reconstruction. C12E8 (n-dodecyl-octaethylene-glycol monoether) was used to fully solubilize various mixtures of lipid and purified Ca2(+)-ATPase, and BioBeads were then used to remove the C12E8. Slow removal resulted in two populations of vesicles, and the proteoliposome population was separated from the liposome population on a sucrose density gradient. These proteoliposomes had a lipid-to-protein ratio of 1:2, and virtually 100% of molecules faced the outside of vesicles, as determined by fluorescein isothiocyanate labeling. Cycles of freeze-thaw caused considerable aggregation of these proteoliposomes, and, if phosphatidyl ethanolamine and phosphatidic acid were included, or if the bilayers were...Continue Reading

Citations

Mar 30, 2010·Journal of Structural and Functional Genomics·Changki KimIban Ubarretxena-Belandia
Jan 16, 1999·Biochimica Et Biophysica Acta·Y HiroakiY Fujiyoshi
Aug 1, 1997·Current Opinion in Structural Biology·J B HeymannA Engel
Nov 18, 1998·Current Opinion in Structural Biology·M H StowellN Unwin
Feb 13, 2001·Micron : the International Research and Review Journal for Microscopy·G Mosser
Jun 19, 2012·The Journal of Biological Chemistry·Delaine K CeholskiHoward S Young
Oct 9, 2013·Proceedings of the National Academy of Sciences of the United States of America·Martin GustavssonGianluigi Veglia
Jan 13, 2015·Science China. Life Sciences·LiGuo Wang, LiGe Tonggu
Sep 3, 1998·Biophysical Journal·J J LacapèreJ L Rigaud
Nov 25, 2000·Biochimica Et Biophysica Acta·J RigaudJ Ranck
Jun 6, 1998·Journal of Structural Biology·L HaslerT Walz
Jun 4, 2016·Journal of Structural Biology·Nicolas CoudrayDavid L Stokes
Jul 20, 2002·Brazilian Journal of Medical and Biological Research = Revista Brasileira De Pesquisas Médicas E Biológicas·J-L Rigaud
Feb 20, 2019·The EMBO Journal·Aljona SitselPoul Nissen
Aug 3, 2019·Proceedings of the National Academy of Sciences of the United States of America·Francesco Tadini-BuoninsegniJens Peter Andersen
Aug 25, 2020·Biochemical Society Transactions·Andrea Marco AmatiChristoph von Ballmoos
Aug 8, 1997·The Journal of Biological Chemistry·G MosserJ L Rigaud
Dec 22, 1999·Annals of the New York Academy of Sciences·H S YoungD L Stokes
May 1, 2001·Journal of Molecular Biology·H S YoungD L Stokes
May 18, 2001·Journal of Structural Biology·M ChamiJ L Rigaud
Feb 22, 2005·Protein Expression and Purification·Jennifer L DouglasHoward S Young
Jul 24, 2021·ChemMedChem·Giacomo SordiFrancesco Tadini-Buoninsegni
Jul 13, 2006·Biochemistry·Michael R AfaraHoward S Young

❮ Previous
Next ❯

Related Concepts

Trending Feeds

COVID-19

Coronaviruses encompass a large family of viruses that cause the common cold as well as more serious diseases, such as the ongoing outbreak of coronavirus disease 2019 (COVID-19; formally known as 2019-nCoV). Coronaviruses can spread from animals to humans; symptoms include fever, cough, shortness of breath, and breathing difficulties; in more severe cases, infection can lead to death. This feed covers recent research on COVID-19.

Blastomycosis

Blastomycosis fungal infections spread through inhaling Blastomyces dermatitidis spores. Discover the latest research on blastomycosis fungal infections here.

Nuclear Pore Complex in ALS/FTD

Alterations in nucleocytoplasmic transport, controlled by the nuclear pore complex, may be involved in the pathomechanism underlying multiple neurodegenerative diseases including Amyotrophic Lateral Sclerosis and Frontotemporal Dementia. Here is the latest research on the nuclear pore complex in ALS and FTD.

Applications of Molecular Barcoding

The concept of molecular barcoding is that each original DNA or RNA molecule is attached to a unique sequence barcode. Sequence reads having different barcodes represent different original molecules, while sequence reads having the same barcode are results of PCR duplication from one original molecule. Discover the latest research on molecular barcoding here.

Chronic Fatigue Syndrome

Chronic fatigue syndrome is a disease characterized by unexplained disabling fatigue; the pathology of which is incompletely understood. Discover the latest research on chronic fatigue syndrome here.

Evolution of Pluripotency

Pluripotency refers to the ability of a cell to develop into three primary germ cell layers of the embryo. This feed focuses on the mechanisms that underlie the evolution of pluripotency. Here is the latest research.

Position Effect Variegation

Position Effect Variagation occurs when a gene is inactivated due to its positioning near heterochromatic regions within a chromosome. Discover the latest research on Position Effect Variagation here.

STING Receptor Agonists

Stimulator of IFN genes (STING) are a group of transmembrane proteins that are involved in the induction of type I interferon that is important in the innate immune response. The stimulation of STING has been an active area of research in the treatment of cancer and infectious diseases. Here is the latest research on STING receptor agonists.

Microbicide

Microbicides are products that can be applied to vaginal or rectal mucosal surfaces with the goal of preventing, or at least significantly reducing, the transmission of sexually transmitted infections. Here is the latest research on microbicides.