Multisite phosphorylation of microtubule-associated protein 2 (MAP-2) in rat brain: peptide mapping distinguishes

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Nuclear Magnetic Resonance Spectroscopy for the Identification of Multiple Phosphorylations of Intrinsically Disordered Proteins
12:47

Nuclear Magnetic Resonance Spectroscopy for the Identification of Multiple Phosphorylations of Intrinsically Disordered Proteins

Published on: December 27, 2016

S I Walaas1, A C Nairn

  • 1Laboratory of Molecular and Cellular Neuroscience, Rockefeller University, New York, New York.

Journal of Molecular Neuroscience : MN
|January 1, 1989
PubMed

Insights

Microtubule-associated protein 2 (MAP-2) undergoes complex phosphorylation by various kinases. Different kinases target distinct sites on MAP-2, influencing its function in neurons.

Area of Science:

  • Neuroscience
  • Molecular Biology
  • Cell Biology

Background:

  • Microtubule-associated protein 2 (MAP-2) is a crucial cytoskeletal protein abundant in neuronal dendrites and perikarya.
  • MAP-2 is known to be regulated by phosphorylation, a process involving various signaling pathways.
  • Understanding MAP-2 phosphorylation is key to elucidating its role in neuronal structure and function.

Purpose of the Study:

  • To investigate the distinct phosphorylation sites on MAP-2 targeted by cyclic AMP-dependent protein kinase, calcium/calmodulin-dependent protein kinase II, and calcium/phospholipid-dependent protein kinase.
  • To compare the phosphorylation patterns of MAP-2 in different regions of the rat brain, specifically the olfactory bulb and cerebral cortex.
  • To characterize the amino acid residues (serine and threonine) involved in MAP-2 phosphorylation by these kinases.

Main Methods:

  • In vitro phosphorylation of MAP-2 using [gamma-32P]ATP with three different purified kinases.
  • Analysis of phosphorylation sites using peptide mapping techniques on MAP-2 isolated from fresh or boiled rat brain cytosol.
  • Comparison of MAP-2 phosphorylation patterns between the olfactory bulb (MAP-2B) and cerebral cortex (MAP-2A and MAP-2B).

Main Results:

  • Cyclic AMP-dependent protein kinase and calcium/calmodulin-dependent protein kinase II commonly phosphorylated four sites on MAP-2, with each kinase also phosphorylating unique sites.
  • Calcium/phospholipid-dependent protein kinase phosphorylated at least 15 sites, with only one site overlapping with the other kinases.
  • Phosphorylation by cyclic AMP-dependent and calcium/phospholipid-dependent protein kinases occurred exclusively on serine residues, while calcium/calmodulin-dependent protein kinase II phosphorylated both serine and threonine residues. No regional differences in peptide maps were observed.

Conclusions:

  • MAP-2 phosphorylation is a complex process involving multiple kinases and a diverse set of phosphorylation sites.
  • The specific phosphorylation pattern of MAP-2 may be regulated by the activity of different kinase pathways.
  • These findings highlight the intricate regulatory mechanisms governing MAP-2 function in situ.

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