Human brain protein phosphorylation in vitro: Cyclic AMP stimulation of electrophoretically-separated substrates

Aryeh Routtenberg*, David G. Morgan, Richard G. Conway, Michael J. Schmidt, Bernardino Ghetti

*Corresponding author for this work

Research output: Contribution to journalArticle

7 Scopus citations

Abstract

In vitro phosphorylation of electrophoretically-separated brain proteins was studied in human frontal cortex obtained 3-16 h post-mortem from 13 patients ages 3 days-82 years with extensive, mild or no neuropathological involvement. In 12 of the 13 cases, cyclic AMP increased incorporation of phosphate into acid-precipitable protein. Analysis of the autoradiographic profiles of separate proteins indicated that phosphorylation of a doublet of molecular weight 86-80,000 was stimulated by cyclic AMP in certain samples. This doublet corresponded to the cyclic AMP stimulated doublet from rat frontal cortex we have termed band D-1,2 (proteins Ia and Ib of Ueda and Greengard20). Of special interest was the fact that, while co-migration was observed in the other phosphoprotein bands studied, band D-1,2 of humans consistently migrated slightly less than rat protein band D-1,2. This difference was not a function of post-mortem time, subcellular fraction or buffer used in the reaction phosphorylation assay. The use of post-mortem tissue was not a contributing factor as the retardation in band D-1,2 migration was still observed when post-mortem rat brain was used for comparison. In two human post-mortem samples, there was no measureable band D-1,2 phosphorylation even in the presence of cyclic AMP. This was the case in both homogenate and crude synaptosome/mitochondrial preparations. Band F-1 (mol. wt. = 47,000) was not observed in any of the human samples studied. This is consistent with prior studies in rat which show that band F-1 phosphorylation is not detected in post-mortem brain. Band F-2 (mol. wt. 41,000) recently identified as pyruvate dehydrogenase11, was lightly phosphorylated under the reaction conditions used in this study.

Original languageEnglish (US)
Pages (from-to)323-333
Number of pages11
JournalBrain research
Volume222
Issue number2
DOIs
StatePublished - Oct 19 1981

Keywords

  • cyclic AMP
  • electrophoresis
  • human brain
  • protein phosphorylation
  • pyruvate dehydrogenase

ASJC Scopus subject areas

  • Neuroscience(all)
  • Molecular Biology
  • Clinical Neurology
  • Developmental Biology

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