Autophosphorylation-dependent protein kinase predominantly phosphorylates Ser115, the in vivo site in brain myelin basic protein

Shiaw-Der Yang, Tze-Jen Huang, Jun-Jae Huang

    Research output: Contribution to journalArticlepeer-review

    3 Citations (Scopus)

    Abstract

    In a previous report [Yang et al., (1987 a), J. Biol Chem.262, 7034-7040], a cyclic-AMP- and calcium-independent brain kinase which requires autophosphorylation for activity was identified as a very potent myelin basic protein (MBP) kinase. In this report, the phosphorylation sites of MBP by this autophosphorylation-dependent protein kinase (autokinase) are further determined by two-dimensional electrophoresis/thin-layer chromatography, phosphoamino acid analysis, high-performance liquid chromatography, tryptic peptide mapping, sequential manual Edman degradation, and direct peptide sequencing. Autokinase phosphorylates MBP on both threonine and serine residues. Three major tryptic phosphopeptide peaks were resolved by C18-reversed phase highper-formance liquid chromatography. Sequential manual Edman degradation together with direct sequence analysis revealed that FS(p)WGAEGQKPGFGYGGR is the phosphorylation site sequence (molar ratio ∼1.0) for the first major phosphopeptide peak. When mapping with bovine brain MBP sequence, we finally demonstrate Ser115, one of the in vivo phosphorylation sites in MBP, as the major site phosphorylated by autokinase, implicating a physiologically relevant role of autokinase in the regulation of brain myelin function. By using the same approach, we also identified HRDT(p)GILDSLGR (molar ratio ∼0.9) and TT(p)HYGSLPQK (molar ratio ∼0.8) as the major phosphorylation site sequences in32P-MBP phosphorylated by autokinase, further indicating that -Arg-XSer/Thr-(neutral amino acid)3-(amino acid-containing hydroxyl group such as Ser/Glu/Asp)-(neutral amino acid)2-may represent a unique consensus sequence motif specifically recognized by this autophosphorylation-dependent multisubstrate/ multifunctional protein kinase in the brain.

    Original languageEnglish
    Pages (from-to)599-607
    Number of pages9
    JournalJournal of Protein Chemistry
    Volume13
    Issue number7
    DOIs
    Publication statusPublished - Oct 1994

    Keywords

    • autophosphorylation-dependent protein serine/threonine kinase
    • consensus sequence motif
    • Myelin basic protein
    • phosphorylation site

    Fingerprint Dive into the research topics of 'Autophosphorylation-dependent protein kinase predominantly phosphorylates Ser<sup>115</sup>, the <i>in vivo</i> site in brain myelin basic protein'. Together they form a unique fingerprint.

    Cite this