Properties of long myosin light chain kinase binding to F-actin in vitro and in vivo

Lula Smith, Mojgan Parizi-Robinson, Min Sheng Zhu, Gang Zhi, Ryosuke Fukui, Kristine E. Kamm, James T. Stull

Research output: Contribution to journalArticle

34 Scopus citations

Abstract

Short and long myosin light chain kinases (MLCKs) are Ca2+/calmodulin-dependent enzymes that phosphorylate the regulatory light chain of myosin II in thick filaments but bind with high affinity to actin thin filaments. Three repeats of a motif made up of the sequence DFRXXL at the N terminus of short MLCK are necessary for actin binding (Smith, L., Su, X., Lin, P., Zhi, G., and Stull, J. T. (1999) J. Biol. Chem. 274, 29433-29438). The long MLCK has two additional DFRXXL motifs and six Ig-like modules in an N-terminal extension, which may confer unique binding properties for cellular localization. Two peptides containing either five or three DFRXXL motifs bound to F-actin and smooth muscle myofilaments with maximal binding stoichiometries consistent with each motif binding to an actin monomer in the filaments. Both peptides cross-linked F-actin and bound to stress fibers in cells. Long MLCK with an internal deletion of the five DFRXXL motifs and the unique NH2-terminal fragment containing six Ig-like motifs showed weak binding. Cell fractionation and extractions with MgCl2 indicate that the long MLCK has a greater affinity for actin-containing filaments than short MLCK in vitro and in vivo. Whereas DFRXXL motifs are necessary and sufficient for short MLCK binding to actin-containing filaments, the DFRXXL motifs and the N-terminal extension of long MLCK confer high affinity binding to stress fibers in cells.

Original languageEnglish (US)
Pages (from-to)35597-35604
Number of pages8
JournalJournal of Biological Chemistry
Volume277
Issue number38
DOIs
StatePublished - Sep 20 2002

ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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