3'-Phosphoadenosine 5'-phosphosulfate biosynthesis and the sulfation of cholecystokinin by the tyrosylprotein-sulfotransferase in rat brain tissue.

This article resumes the work we have accomplished in the past few years. Cholecystokinin sulfation is an important post-translational modification necessary for the biological activity of this peptide hormone. The tyrosyl protein sulfotransferase (TPST) activity from rat cerebral cortex was characterized.

Subcellular and developmental studies of the tyrosyl protein sulfotransferase in rat brain.

1. Tyrosyl protein sulfotransferase (TPS) activity in the newborn and mature rat brain was studied using the cholecystokinin derivative terbutyloxycarbonyl-Asp-Tyr-Met-Gly-Trp-Met-Asp-PheNH2, BocCCK-8(ns), as the peptide substrate. 2.

Cholecystokinin activation: evidence for an ordered reaction mechanism for the tyrosyl protein sulfotransferase responsible for the peptide sulfation.

The kinetics of the forward tyrosyl protein sulfotransferase (TPS) reaction were examined using an assay based on the 35SO4 transfer from 3'-phosphoadenosine 5'-phospho(35S)sulfate [( 35S]PAPS) to tyrosyl residues of the non-sulfated cholecystokinin derivative, BocCCK-8(ns). TPS present in the microsomal membranes from rat cerebral cortex was used for these studies. Initial velocity measurements performed over a wide range of PAPS, BocCCK-8(ns), 3'-PAP and BocCCK-8(s) concentrations, indicated that the reaction follows an ordered mechanistic pathway.

Characterization of a tyrosine sulfotransferase in rat brain using cholecystokinin derivatives as acceptors.

An apparently novel tyrosyl sulfotransferase activity was detected in a crude microsomal fraction from rat cerebral cortex by using 3'-phosphoadenosine 5'-phospho[35S]sulfate [( 35S]PAPS) as the sulfate donor and various cholecystokinin (CCK) fragments or derivatives as acceptors. Among the latter, the shortest substrate was tert-butoxycarbonylaspartyltyrosine (Boc-Asp-Tyr), but the reaction was optimized by increasing the length of the peptide sequence on the C-terminal side up to tert-butoxycarbonylcholecystokinin octapeptide (Boc-CCK-8) as well as by the presence of acidic amino acid residues at the N-terminal side. Peptides with an N-terminal Tyr residue (e.

Sulfation and desulfation of cerebral cholecystokinin.

An apparently novel sulfotransferase present in microsomal and vesicular fractions from rat brain is able to transfer [35S]sulfate groups from [35S]PAPS to CCK derivatives. Its optimum pH (approximately 6), its substrate specificity, and its subcellular localization are all consistent with its function as post-translational processing enzyme. However, its presence in tissues devoid of CCK argues against this idea, or could simply mean that it is involved in the processing of other peptides besides CCK.