2-color photobleaching experiments reveal distinct intracellular dynamics of two components of the Hsp90 complex

Autor: Elena Suslova, Didier Picard, Pierre-André Briand
Jazyk: angličtina
Rok vydání: 2006
Předmět:
Cytosol/metabolism
congenital
hereditary
and neonatal diseases and abnormalities
Molecular Chaperones/chemistry/genetics/metabolism
Recombinant Fusion Proteins
Green Fluorescent Proteins
Biology
Transfection
HSP90 Heat-Shock Proteins/chemistry/genetics/metabolism
Green fluorescent protein
chemistry.chemical_compound
Cytosol
Subcellular Fractions/metabolism
ddc:570
Mitochondria/metabolism
polycyclic compounds
Humans
HSP90 Heat-Shock Proteins
skin and connective tissue diseases
Prostaglandin-E Synthases
Fluorescent Dyes
Green Fluorescent Proteins/chemistry/genetics/metabolism
Phosphoproteins/chemistry/genetics/metabolism
Recombinant Fusion Proteins/chemistry/genetics/metabolism
Fluorescence recovery after photobleaching
Cell Biology
Geldanamycin
Phosphoproteins
Fusion protein
Photobleaching
Hsp90
Mitochondria
Cell biology
chemistry
Hela Cells
Multiprotein Complexes
biology.protein
mCherry
HeLa Cells
Molecular Chaperones
Subcellular Fractions
Fluorescence Recovery After Photobleaching
Zdroj: Experimental Cell Research, Vol. 312, No 19 (2006) pp. 3949-58
ISSN: 0014-4827
Popis: The abundant molecular chaperone Hsp90 functions in association with co-chaperones including p23 to promote the folding and maturation of a subset of cytosolic proteins. "Fluorescence recovery after photobleaching" (FRAP) experiments showed that the dynamics of p23 in live cells is dictated by Hsp90. Since Hsp90 is present in large excess over p23, the mobility of Hsp90 could conceivably be quite different. To facilitate the analysis and to allow a direct comparison with p23, we developed a 2-color FRAP technique. Two test proteins are expressed as fusion proteins with the two spectrally separable fluorescent proteins mCherry and enhanced green fluorescent protein (EGFP). The 2-color FRAP technique is powerful for the concomitant recording of two proteins located in the same area of a cell, two components of the same protein complex, or mutant and wild-type versions of the same protein under identical experimental conditions. 2-color FRAP of Hsp90 and p23 is virtually indistinguishable, consistent with the notion that they are both engaged in a multitude of large protein complexes. However, when Hsp90-p23 complexes are disrupted by the Hsp90 inhibitor geldanamycin, p23 moves by free diffusion while Hsp90 maintains its low mobility because it remains bound in remodeled multicomponent complexes.
Databáze: OpenAIRE
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