Open AccessCharacterization of a calcium‐dependent calmodulin‐binding domain in the 135‐kD human protein 4.1 isoform
Author(s) -
Leclerc Estelle,
Vetter Stefan
Publication year - 1998
Publication title -
european journal of biochemistry
Language(s) - English
Resource type - Journals
eISSN - 1432-1033
pISSN - 0014-2956
DOI - 10.1046/j.1432-1327.1998.2580567.x
Subject(s) - calmodulin , peptide , phosphodiesterase , protein kinase a , phosphorylation , target peptide , chemistry , biochemistry , gene isoform , calcium , n terminus , binding protein , kinase , binding site , peptide sequence , microbiology and biotechnology , enzyme , biology , gene , organic chemistry
The putative calmodulin binding domain of non‐erythroid protein 4.1, previously suggested by Kelly et al. [Kelly, G. M., Zelus, B. D. & Moon, R. T. (1991) J. Biol. Chem. 266 , 12 469−12 473] has been synthesized, and its binding to calmodulin has been studied by fluorescence spectroscopy. For this purpose, the peptide has been N‐terminally dansylated. The 4.1 peptide Dns‐Abu‐S 76 RGLSRLFSSFLKRPKS 92 , binds calmodulin in a calcium‐dependent way with high affinity ( K d = 23 ± 6 nM). The peptide inhibits bovine‐heart phosphodiesterase with an IC 50 of 50 nM. Since the sequence of the peptide shows two putative consensus sites of phosphorylation by cAMP‐dependent protein kinase or Ca 2+ −calmodulin protein‐kinase II, the interaction of the two mono‐phosphorylated peptides (P4.1 Ser 80‐P and P4.1 Ser 84‐P ) and the di‐phosphorylated peptide (P4.1 Ser 80‐P /Ser 84‐P ) with calmodulin has been investigated. A decrease of affinity by a factor 1.5−8 has been observed for the phosphorylated peptides. CD measurements have shown an increase of the content of α helices in the peptides when bound to calmodulin.