Visualization of inositol 1,4,5-trisphosphate receptor by atomic force microscopy.
Neurosci Lett
; 391(3): 102-7, 2006 Jan 02.
Article
em En
| MEDLINE
| ID: mdl-16198054
ABSTRACT
Inositol 1,4,5-trisphosphate (IP(3)) receptor (IP(3)R) acts as a ligand-gated channel that mediates neuronal signals by releasing Ca(2+) from the endoplasmic reticulum. The three-dimensional (3D) structure of tetrameric IP(3)R has been demonstrated by using electron microscopy (EM) with static specimens; however, the dynamic aspects of the IP(3)R structure have never been visualized in a native environment. Here we attempt to measure the surface topography of IP(3)R in solution using atomic force microscopy (AFM). AFM revealed large protrusions extending approximately 4.3 nm above a flat membrane prepared from Spodoptera frugiperda (Sf9) cells overexpressing mouse type 1 IP(3)R (Sf9-IP(3)R1). The average diameter of the large protrusions was approximately 32 nm. A specific antibody against a cytosolic epitope close to the IP(3)-binding site enabled us to gold-label the Sf9-IP(3)R1 membrane as confirmed by EM. AFM images of the gold-labeled membrane revealed 7.7-nm high protrusions with a diameter of approximately 30 nm, which should be IP(3)R1-antibody complexes. Authentic IP(3)R1 immuno-purified from mouse cerebella had approximately the same dimensions as those of the IP(3)R-like protrusions on the membrane. Altogether, these results suggest that the large protrusions on the Sf9-IP(3)R1 membrane correspond to the cytosolic domain of IP(3)R1. Our study provides the first 3D representation of individual IP(3)R1 particles in an aqueous solution.
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Base de dados:
MEDLINE
Assunto principal:
Água
/
Canais de Cálcio
/
Receptores Citoplasmáticos e Nucleares
/
Microscopia de Força Atômica
Idioma:
En
Ano de publicação:
2006
Tipo de documento:
Article