Purification of plasma membrane and extraction of channels from gastric mucosa

Academic Article

Abstract

  • Large scale purification of membrane fractions of dog gastric mucosa was achieved using zonal density gradient fractionation of the 100,000g fraction of mucosal homogenates. The criteria of purity used were EM appearance, enrichment of HCO3- ATPase and AMPase activity and absence of MAO, SDH, β glucuronidase and RNA. In linear sucrose gradients, a single band of two types of membrane fragments (vesicles and sheets) was observed at 29% sucrose (w/w). In ficoll sucrose gradients two membrane peaks were obtained, free of MAO or SDH activity, the lighter fraction having a much higher AMPase activity. Using the 8,000g precipitate, purified gastric mitochondria were also obtained. Triton X 100 extracts of these fractions were tested for ionophore activity in chambered bilayer preparations. Only the membrane fraction extract showed activity. At least three types of channel forming activity were found, based on channel conductance and ion selectivity. These membrane preparations are, therefore, suitable for study of gastric transport mechanisms.
  • Authors

    Published In

  • The FASEB Journal  Journal
  • Author List

  • Spenney JG; Goodall MC; Sachs G
  • Start Page

  • 986
  • Volume

  • 32
  • Issue

  • 3 I