Purification of synaptic vesicles from elasmobranch electric organ and the use of biophysical criteria to demonstrate purity. Academic Article uri icon

Overview

abstract

  • We have purified cholinergic synaptic vesicles from the electric organs of two related marine elasmobranchs, Torpedo californica and Narcine brasiliensis, to a specific activity higher than had previously been obtained. We have demonstrated the homogeneity of the vesicles by biophysical criteria. The purification scheme consisted of differential centrifugation, flotation equilibrium in sucrose density gradients, and permeation chromatography on glass bead columns of average pore size 3000 A. Our criteria for purity were that bound acetylcholine, bound nucleotide triphosphate, protein, and lipid--phosphorus behave identically when vesicles were analyzed by procedures which depend on vesicle size, density, and charge. Contaminants were not detected when vesicles were fractionated by preparative and analytical sedimentation, by preparative equilibrium sedimentation using glycerol density gradients, or by electrophoresis in Ficoll density gradients. Pure synaptic vesicles, which have been purified 290-fold from the initial homogenate, contain per mg of protein: 8 mumol of acetylcholine, 3 mumol of ATP, and 7 mumol of lipid phosphorus. These procedures may be of general value in the purification of membrane vesicles.

publication date

  • April 4, 1978

Research

keywords

  • Electric Organ
  • Synaptic Vesicles

Identity

Scopus Document Identifier

  • 0017821107

Digital Object Identifier (DOI)

  • 10.1021/bi00600a009

PubMed ID

  • 418798

Additional Document Info

volume

  • 17

issue

  • 7