Phosphoproteins of the Adrenal Chromaffin Granule Membrane

Abstract

A fraction of chromaffin granule membranes contained a number of substrates for endogenous protein kinase activity as well as endogenous phosphatase activity. The major 32P-labelled polypeptide of molecular weight 43,000 appeared to be the alpha-subunit of pyruvate dehydrogenase of residual mitochondria. Several polypeptides showed cyclic AMP stimulation of phosphorylation of which the major polypeptide of molecular weight 59,000 shows half-maximal phosphorylation with 0.49 microM cyclic AMP. The phosphorylation of several other polypeptides is inhibited at high cyclic AMP concentrations. From studies with immunoprecipitation and two-dimensional electrophoresis it was found that alpha- and beta-tubulin and actin were absent from the granule membranes. However 32P labelling of a proportion of the copies of dopamine-beta-hydroxylase was demonstrated. The majority of the substrates for endogenous protein kinase activity are probably on the cytoplasmic side of the granule membrane.

Exocytosis induction in Paramecium tetraurelia cells by exogenous phosphoprotein phosphatase in vivo and in vitro: possible involvement of calcineurin in exocytotic membrane fusion.

“…The probable involvement of a Ca2+-CaM-CaN complex in exocytosis induction via dephosphorylation of the 65-kD PP can also be inferred from experiments with isolated cortices (Figs. [5][6][7][8][9]. For these preparations Vilmart-Seuwen et al (65) have documented that the membranes of trichocysts attached to such cortices fuse perfectly with the cell membrane and that trichocyst ghosts do not reseal.…”

Rat dopamine β‐hydroxylase: Molecular cloning and characterization of the cDNA and regulation of the mRNA by reserpine

“…At present it is uncertain if this phosphorylation site is utilized. There is a report of low levels of phosphorylation of DBH in bovine adrenal chromaffin cells (Burgoyne and Geisow, 1982). However, attempts to label DBH in PC12 cells with radioactive phosphate failed to detect any phosphorylated DBH under conditions in which tyrosine hydroxylase was extensively phosphorylated (Sabban et al, 1983a;McHugh et al, 1985).…”

The primary structure of human dopamine-beta-hydroxylase: insights into the relationship between the soluble and the membrane-bound forms of the enzyme.

“…The exposure time was 3 h. 1981) are found, indicating that glycosylation is limited by the availability of some of these sites to the glycosylating enzymes. A potential phosphorylation site for calmodulin-dependent kinase is also present (Pearson et al, 1985); interestingly, it overlaps with one of the possible glycosylation sites, a situation that might explain conflicting reports on the phosphorylation of mature DBH (Burgoyne and Geisow, 1982;Sabban et al, 1983;McHugh et al, 1985).…”