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Arch Biochem Biophys. 2000 May 1;377(1):80-4.

Determination of a cAMP-dependent protein kinase phosphorylation site in the C-terminal region of human endothelial actin-binding protein.

Author information

1
Departamento de Cultivo de Tejidos, Instituto Nacional de Cardiología, Ignacio Chávez, Juan Badiano #1, D. F. México. jay@mailer.main.conacyt.mx

Abstract

Three different C-terminal regions of human endothelial actin-binding protein-280 (ABP-280 or ABP; nonmuscle filamin) were subcloned and efficiently expressed in the Escherichia coli BL21 (DE3) system as indicated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. As predicted by the aminoacid sequence one of the fragments, a 109-kDa peptide (residues 1671-2647), contained a calpain cleavage site and two potential cAMP-dependent protein kinase (PKA) phosphorylation sites (serine 2152 and threonine 2336). A second fragment, a 74-kDa peptide (residues 1671-2331), contained a calpain cleavage site and one of the three presumptive PKA phosphorylation sites (serine 2152). The third fragment, a 48-kDa peptide (residues 2223-2647), contained only one of the PKA sites (threonine 2336). Phosphorylation of these truncated peptides indicated that only the fragments containing serine 2152 incorporated phosphate after PKA treatment. Site-directed mutagenesis analysis confirmed that serine 2152 is the unique substrate for PKA in the C-terminal region of ABP. The functional significance of phosphorylation of this residue, which belongs to a serine-proline motif, is discussed.

PMID:
10775444
DOI:
10.1006/abbi.2000.1762
[Indexed for MEDLINE]

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