Quantitative measurements of Ca2+/calmodulin binding and activation of myosin light chain kinase in cells

Ramaz Geguchadze, Gang Zhi, Kim S. Lau, Eiji Isotani, Anthony Persechini, Kristine E. Kamm, James T. Stull

Research output: Contribution to journalArticle

37 Citations (Scopus)

Abstract

Myosin II regulatory light chain (RLC) phosphorylation by Ca 2+/calmodulin (CaM)-dependent myosin light chain kinase (MLCK) is implicated in many cellular actin cytoskeletal functions. We examined MLCK activation quantitatively with a fluorescent biosensor MLCK where Ca 2+-dependent increases in kinase activity were coincident with decreases in fluorescence resonance energy transfer (FRET) in vitro. In cells stably transfected with CaM sensor MLCK, increasing [Ca2+] i increased MLCK activation and RLC phosphorylation coincidently. There was no evidence for CaM binding but not activating MLCK at low [Ca 2+]i. At saturating [Ca2+]i MLCK was not fully activated probably due to limited availability of cellular Ca 2+/CaM.

Original languageEnglish (US)
Pages (from-to)121-124
Number of pages4
JournalFEBS Letters
Volume557
Issue number1-3
DOIs
StatePublished - Jan 16 2004

Fingerprint

Myosin-Light-Chain Kinase
Calmodulin
Chemical activation
Phosphorylation
Myosin Type II
Myosin Light Chains
Fluorescence Resonance Energy Transfer
Biosensing Techniques
Biosensors
Actins
Phosphotransferases
Availability
Light
Sensors

Keywords

  • Calcium
  • Calmodulin
  • Fluorescence resonance energy transfer
  • Myosin light chain kinase
  • Phosphorylation

ASJC Scopus subject areas

  • Biochemistry
  • Biophysics
  • Molecular Biology

Cite this

Quantitative measurements of Ca2+/calmodulin binding and activation of myosin light chain kinase in cells. / Geguchadze, Ramaz; Zhi, Gang; Lau, Kim S.; Isotani, Eiji; Persechini, Anthony; Kamm, Kristine E.; Stull, James T.

In: FEBS Letters, Vol. 557, No. 1-3, 16.01.2004, p. 121-124.

Research output: Contribution to journalArticle

Geguchadze, Ramaz ; Zhi, Gang ; Lau, Kim S. ; Isotani, Eiji ; Persechini, Anthony ; Kamm, Kristine E. ; Stull, James T. / Quantitative measurements of Ca2+/calmodulin binding and activation of myosin light chain kinase in cells. In: FEBS Letters. 2004 ; Vol. 557, No. 1-3. pp. 121-124.
@article{d6832d160a7d4efdb2ca58403ca1eb03,
title = "Quantitative measurements of Ca2+/calmodulin binding and activation of myosin light chain kinase in cells",
abstract = "Myosin II regulatory light chain (RLC) phosphorylation by Ca 2+/calmodulin (CaM)-dependent myosin light chain kinase (MLCK) is implicated in many cellular actin cytoskeletal functions. We examined MLCK activation quantitatively with a fluorescent biosensor MLCK where Ca 2+-dependent increases in kinase activity were coincident with decreases in fluorescence resonance energy transfer (FRET) in vitro. In cells stably transfected with CaM sensor MLCK, increasing [Ca2+] i increased MLCK activation and RLC phosphorylation coincidently. There was no evidence for CaM binding but not activating MLCK at low [Ca 2+]i. At saturating [Ca2+]i MLCK was not fully activated probably due to limited availability of cellular Ca 2+/CaM.",
keywords = "Calcium, Calmodulin, Fluorescence resonance energy transfer, Myosin light chain kinase, Phosphorylation",
author = "Ramaz Geguchadze and Gang Zhi and Lau, {Kim S.} and Eiji Isotani and Anthony Persechini and Kamm, {Kristine E.} and Stull, {James T.}",
year = "2004",
month = "1",
day = "16",
doi = "10.1016/S0014-5793(03)01456-X",
language = "English (US)",
volume = "557",
pages = "121--124",
journal = "FEBS Letters",
issn = "0014-5793",
publisher = "Elsevier",
number = "1-3",

}

TY - JOUR

T1 - Quantitative measurements of Ca2+/calmodulin binding and activation of myosin light chain kinase in cells

AU - Geguchadze, Ramaz

AU - Zhi, Gang

AU - Lau, Kim S.

AU - Isotani, Eiji

AU - Persechini, Anthony

AU - Kamm, Kristine E.

AU - Stull, James T.

PY - 2004/1/16

Y1 - 2004/1/16

N2 - Myosin II regulatory light chain (RLC) phosphorylation by Ca 2+/calmodulin (CaM)-dependent myosin light chain kinase (MLCK) is implicated in many cellular actin cytoskeletal functions. We examined MLCK activation quantitatively with a fluorescent biosensor MLCK where Ca 2+-dependent increases in kinase activity were coincident with decreases in fluorescence resonance energy transfer (FRET) in vitro. In cells stably transfected with CaM sensor MLCK, increasing [Ca2+] i increased MLCK activation and RLC phosphorylation coincidently. There was no evidence for CaM binding but not activating MLCK at low [Ca 2+]i. At saturating [Ca2+]i MLCK was not fully activated probably due to limited availability of cellular Ca 2+/CaM.

AB - Myosin II regulatory light chain (RLC) phosphorylation by Ca 2+/calmodulin (CaM)-dependent myosin light chain kinase (MLCK) is implicated in many cellular actin cytoskeletal functions. We examined MLCK activation quantitatively with a fluorescent biosensor MLCK where Ca 2+-dependent increases in kinase activity were coincident with decreases in fluorescence resonance energy transfer (FRET) in vitro. In cells stably transfected with CaM sensor MLCK, increasing [Ca2+] i increased MLCK activation and RLC phosphorylation coincidently. There was no evidence for CaM binding but not activating MLCK at low [Ca 2+]i. At saturating [Ca2+]i MLCK was not fully activated probably due to limited availability of cellular Ca 2+/CaM.

KW - Calcium

KW - Calmodulin

KW - Fluorescence resonance energy transfer

KW - Myosin light chain kinase

KW - Phosphorylation

UR - http://www.scopus.com/inward/record.url?scp=1642532373&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=1642532373&partnerID=8YFLogxK

U2 - 10.1016/S0014-5793(03)01456-X

DO - 10.1016/S0014-5793(03)01456-X

M3 - Article

C2 - 14741352

AN - SCOPUS:1642532373

VL - 557

SP - 121

EP - 124

JO - FEBS Letters

JF - FEBS Letters

SN - 0014-5793

IS - 1-3

ER -