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Effects of solutes on the formation of crystalline sheets of the Ca(2+)-ATPase in detergent-solubilized sarcoplasmic reticulum.
- Source :
-
Biochimica et biophysica acta [Biochim Biophys Acta] 1991 Dec 09; Vol. 1070 (2), pp. 374-86. - Publication Year :
- 1991
-
Abstract
- The Ca(2+)-ATPase crystals formed in detergent solubilized sarcoplasmic reticulum (SR) at 2 degrees C in a crystallization medium of 0.1 M KCl, 10 mM K-Mops (pH 6.0), 3 mM MgCl2, 3 mM NaN3, 5 mM DTT, 25 IU/ml Trasylol, 2 micrograms/ml 1,6-di-tert-butyl-p-cresol, 20% glycerol and 20 mM CaCl2 (J. Biol. Chem. 263, 5277 and 5287 (1988)) contain highly ordered sheets of ATPase molecules, that associate into large multilamellar stacks (greater than 100 layers). When the crystallization is performed in the same medium but in the presence of 40% glycerol at low temperature the stacking is reduced to 4-5 layers and the average diameter of the crystalline sheets is increased from less than 1 micron to 2-3 microns. Glycerol and low temperature presumably reduce stacking by interfering with the interactions between the hydrophilic headgroups of Ca(2+)-ATPase molecules in adjacent lamellae, while not affecting or promoting the ordering of ATPase molecules within the individual sheets. Electron diffraction patterns could be regularly obtained at 8 A and occasionally at 7 A resolution on crystals formed in 40% glycerol, either at 2 degrees C or at -70 degrees C. In the same media but in the absence of glycerol, polyethyleneglycol 1450, 3000 and 8000 (1-8%) induced the formation of ordered crystalline arrays containing 10-12 layers that were similar to those obtained in 40% glycerol. Replacement of 40% glycerol with 10-50% glucose or supplementation of the standard crystallization medium with polyethyleneglycol (PEG 3000 or 8000; 1, 2, 5 and 8%) had no beneficial effect on the order of crystalline arrays compared with media containing 40% glycerol.
- Subjects :
- Animals
Calcium-Transporting ATPases chemistry
Crystallization
Detergents
Microscopy, Electron
Muscles enzymology
Protein Conformation
Rabbits
Sarcoplasmic Reticulum ultrastructure
Solvents
Calcium-Transporting ATPases metabolism
Calcium-Transporting ATPases ultrastructure
Sarcoplasmic Reticulum enzymology
Subjects
Details
- Language :
- English
- ISSN :
- 0006-3002
- Volume :
- 1070
- Issue :
- 2
- Database :
- MEDLINE
- Journal :
- Biochimica et biophysica acta
- Publication Type :
- Academic Journal
- Accession number :
- 1837235
- Full Text :
- https://doi.org/10.1016/0005-2736(91)90078-m