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Preparation of a fluorescent-labeled der ... for calmodulin binding proteins
Preparation of a fluorescent-labeled derivative of calmodulin which retains its affinity for calmodulin binding proteins.
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Calmodulin was derivatized with 5-[[[(iodoacetyl)amino]ethyl]amino]-1-naphthalenesulfonic acid to fluorescently label the protein. This derivative (AEDANS.CaM) stimulated the Ca2+-sensitive cyclic nucleotide phosphodiesterase and formed Ca2+-dependent complexes with troponin I and the phosphodiesterase. Association between AEDANS.CaM and these proteins was directly monitored by changes in fluorescence anisotropy. The dissociation constants for the AEDANS.CaM--troponin I and AEDANS.CaM--phosphodiesterase complexes were 60 nM and 4 nM, respectively. This fluorescent derivative of calmodulin appears suitable for direct monitoring of the complexes between calmodulin and calmodulin binding proteins. Rotational diffusion of AEDANS.CaM was also measured with fluorescence anisotropy. These measurements indicated that the shape of calmodulin in solution is best approximated by a prolate ellipsoid.
LaPorte DC, Keller CH, Olwin BB, Storm DR
Biochemistry
1981-07-07 00:00
20
14
3965-72
Animals,Brain,Calcium,Calcium-Binding Proteins,Calmodulin,Calmodulin-Binding Proteins,Carrier Proteins,Cattle,Fluorescence Polarization,Fluorescent Dyes,Kinetics,Naphthalenesulfonates,Phosphoric Diester Hydrolases,Protein Binding,Protein Conformation,AEDANS-calmodulin,Calcium-Binding Proteins,Calmodulin,Calmodulin-Binding Proteins,Carrier Proteins,Fluorescent Dyes,Naphthalenesulfonates,Calcium,Phosphoric Diester Hydrolases
Biochemistry
NIAID AI 00120, NHLBI HL 05933, NHLBI HL 23606
0006-2960
884
True
6269577
