Tryptophan residues in caldesmon are major determinants for calmodulin binding

Biochemistry
S P GraetherA S Mak

Abstract

Calmodulin has been shown to interact with the COOH-terminal domain of gizzard h-caldesmon at three sites, A (residues 658-666), B (residues 687-695), and B' (residues 717-725), each of which contains a Trp residue [Zhan et al. (1991) J. Biol. Chem. 266, 21810-21814; Marston et al. (1994) J. Biol. Chem. 296, 8134-8139; Mezgueldi et al. (1994) J. Biol. Chem. 269, 12824-12832]. To determine the contribution of each of the three Trp residues in the calmodulin-caldesmon interaction, we have mutated the Trp residues to Ala in the COOH-terminal domain of fibroblast caldesmon (CaD39) and studied the effects on calmodulin binding by fluorescence measurements and using immobilized calmodulin. Wild-type CaD39 binds with a Kd of 0.13 x 10(-6) M and a stoichiometry of 1 mol of calmodulin per mol of caldesmon. Replacing Trp 659 at site A or Trp 692 at site B to Ala reduces binding by 22- and 31-fold (Kd = 2.9 x 10(-6) and 4.0 x 10(-6) M), respectively, and destabilizes the CaD39-calmodulin complex by 1.75 and 1.94 kcal mol-1, respectively. Mutation of both Trp 659 and Trp 692 to Ala further reduces binding with a Kd of 6.1 x 10(-6) M and destabilizes the complex by 2.17 kcal mol-1. On the other hand, mutation of Trp 722 at site B' to Ala ca...Continue Reading

References

Oct 1, 1991·The Biochemical Journal·S B Marston, C S Redwood
Aug 1, 1991·Journal of Muscle Research and Cell Motility·J Bryan, R Lee
Feb 1, 1990·Trends in Biochemical Sciences·K T O'Neil, W F DeGrado
Dec 1, 1985·Journal of Muscle Research and Cell Motility·S B Marston, C W Smith
Aug 1, 1995·The Biochemical Journal·T S TsurudaA S Mak
Feb 1, 1993·Current Opinion in Cell Biology·F Matsumura, S Yamashiro
Jun 1, 1996·The Biochemical Journal·P A HuberS B Marston

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Citations

May 9, 2001·Biochimica Et Biophysica Acta·M V MedvedevaN B Gusev
Mar 7, 1998·FEBS Letters·A A PolyakovN B Gusev
Apr 8, 2009·International Review of Cell and Molecular Biology·Jim Jung-Ching LinJian-Ping Jin
Apr 6, 2006·Journal of Cell Science·Robert EvesAlan S Mak

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