Purification of type 1 protein (serine/threonine) phosphatases by microcystin-Sepharose affinity chromatography

Greg Moorhead, Robert W. MacKintosh, Nick Morrice, Timothy Gallagher, Carol MacKintosh

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    151 Citations (Scopus)

    Abstract

    A microcystin (MC)-Sepharose column was prepared by addition of 2-aminoethanethiol to the a,ß-unsaturated carbonyl of the N-methyldehydroalanine residue of MC-LR, followed by reaction of the introduced amino group with N-hydroxysuccinimide-activated CH-Sepharose. The MC-Sepharose bound protein phosphatase-1 (PP1) with high capacity and purified human PP1? in one step from E. coli extracts. It was also used to purify forms of PP1 bound to myofibrils from skeletal muscle. Two of these comprised PP1 complexed to N-terminal fragments of the M-subunit which enhance its myosin phosphatase activity, while the third comprised PP1 and an N-terminal fragment of the glycogen-binding (G)-subunit.
    Original languageEnglish
    Pages (from-to)46-50
    Number of pages5
    JournalFEBS Letters
    Volume356
    Issue number1
    DOIs
    Publication statusPublished - 1 Dec 1994

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