English
 
User Manual Privacy Policy Disclaimer Contact us
  Advanced SearchBrowse

Item

ITEM ACTIONSEXPORT

Released

Journal Article

The separation and characterization of two forms of Torpedo electric organ calelectrin.

MPS-Authors
/persons/resource/persons15095

Fritsche,  U.
Abteilung Neurochemie, MPI for biophysical chemistry, Max Planck Society;

/persons/resource/persons15662

Potschka,  M.
Department of Biochemistry and Cell Biology, MPI for biophysical chemistry, Max Planck Society;

/persons/resource/persons16015

Whittaker,  V. P.
Abteilung Neurochemie, MPI for biophysical chemistry, Max Planck Society;

Locator
There are no locators available
Fulltext (public)
There are no public fulltexts available
Supplementary Material (public)
There is no public supplementary material available
Citation

Fritsche, U., von Kieckebusche, A., Potschka, M., Whittaker, V. P., & Witzemann, V. (1988). The separation and characterization of two forms of Torpedo electric organ calelectrin. Biochimica et Biophysica Acta-Protein Structure and Molecular Enzymology, 957(1), 122-130. doi:10.1016/0167-4838(88)90164-1.


Cite as: http://hdl.handle.net/11858/00-001M-0000-002D-2F4D-1
Abstract
Two methods for extracting calelectrin, a Ca2+-regulated membrane-binding protein from the electric organ ofTorpedo marmorata, have been compared and the more promising one was modified to increase the yield to 7–8 mg · kg−1 wet weight of tissue, that is 4–5-times greater than the original method. The calelectrin so obtained could be resolved into a minor component (designated L-calelectrin) eluted from an anion-exchange column at relatively low ionic strength (100 mM NaCl) and a major component (H-calelectrin) eluted at higher ionic strength (300 mM NaCl). The two forms were also separated by chromatography on a hydrophobic resin. Electrophoresis on cellulose acetate indicated that L-calelectrin had a lower mean isoelectric point that the H-form and polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulphate showed that under reducing conditions (presence of 5% β-mercaptoethanol) both forms migrated as single species, the L-form having a lower apparent relative molecular mass (Mr 32 000) than the H-form (34 000). Under non-reducing conditions, there was no change in the migration of L-calelectrin but the H-form was resolved into two components of Mr 34 000 and 32 000. The addition of 2 mM Ca2+ had no effect on the migration of either form. Both forms were equally recognized by an anti-calelectrin antiserum and were microheterogeneous with respect to their isoelectric points (pH 4.3–5.5) in two-dimensional gel electrophoresis. Physical measurements were carried out on the major H-form. The Stokes radius was estimated to be 3 nm, corresponding to an apparent Mr of 44 000. It was unaffected by changes in ionic strength, pH or Ca2+ concentration. Analytical ultracentrifugation gave a sedimentation constant of 2.9 S and an apparent Mr of 36 000. Measurements of circular dichroism indicated that 78% of the molecule was in the α-helix configuration and 22% in random coil. Ca2+ had no significant effect on the conformation.