Medizinische Universität Graz Austria/Österreich - Forschungsportal - Medical University of Graz

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"Köpfe" der aktuellen Meldungen:

Nina Höller

Nariae Baik-Schneditz

Bernhard Schwaberger

Lukas Peter Mileder

Niels Hammer

Gerhard Pichler

Roland Malli

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Osibow, K; Malli, R; Kostner, GM; Graier, WF.
A new type of non-Ca2+-buffering Apo(a)-based fluorescent indicator for intraluminal Ca2+ in the endoplasmic reticulum.
J BIOL CHEM. 2006; 281(8): 5017-5025. Doi: 10.1074/jbc.M508583200 [OPEN ACCESS]
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Führende Autor*innen der Med Uni Graz: Graier Wolfgang; Osibow Karin
Co-Autor*innen der Med Uni Graz: Kostner Gerhard; Malli Roland
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Abstract:: Genetically encoded Ca2+ indicators are outstanding tools for the assessment of intracellular/organelle Ca2+ dynamics. Basically, most indicators contain the Ca2+-binding site of a (mutated) cytosolic protein that interacts with its natural (mutated) interaction partner upon binding of Ca2+. Consequently, a change in the structure of the sensor occurs that, in turn, alters the fluorescent properties of the sensor. Herein, we present a new type of genetically encoded Ca2+ indicator for the endoplasmic reticulum (ER) (apoK1-er (W. F. Graier, K. Osibow, R. Malli, and G. M. Kostner, patent application number 05450006.1 at the European patent office)) that is based on a single kringle domain from apolipoprotein(a), which is flanked by yellow and cyan fluorescent protein at the 3'- and 5'-ends, respectively. Notably, apoK1-er does not interact with Ca2+ itself but serves as a substrate for calreticulin, the main constitutive Ca2+-binding protein in the ER. ApoK1-er assembles with calreticulin and the protein disulfide isomerase ERp57 and undergoes a conformational shift in a Ca2+-dependent manner that allows fluorescence resonance energy transfer between the two fluorophores. This construct primarily offers three major advantages compared with the already existing probes: (i) it resolves perfectly the physiological range of the free Ca2+ concentration in the ER, (ii) expression of apoK1-er does not affect the Ca2+ buffering capacity of the ER, and (iii) apoK1-er is not inactivated by binding of constitutive interaction partners that prevent Ca2+-dependent conformational changes. These unique characteristics of apoK1-er make this sensor particularly attractive for studies on ER Ca2+ signaling and dynamics in which alteration of Ca2+ fluctuations by expression of any additional Ca2+ buffer essentially has to be avoided.
Find related publications in this database (using NLM MeSH Indexing): Apolipoproteins A - chemistry; Binding Sites - chemistry; Buffers - chemistry; Calcium - metabolism; Calreticulin - metabolism; Cell Line - metabolism; Cytosol - metabolism; Detergents - pharmacology; Endoplasmic Reticulum - metabolism; Fluorescence Resonance Energy Transfer - metabolism; Fluorescent Dyes - pharmacology; Heat-Shock Proteins - metabolism; Homeostasis - metabolism; Humans - metabolism; Hydrogen-Ion Concentration - metabolism; Immunohistochemistry - metabolism; Immunoprecipitation - metabolism; Mutation - metabolism; Oxidation-Reduction - metabolism; Protein Binding - metabolism; Protein Conformation - metabolism; Protein Disulfide-Isomerase - metabolism; Protein Structure, Tertiary - metabolism; Research Support, Non-U.S. Gov't - metabolism; Time Factors - metabolism