PubMed Journals: Biochem J

  Source:		PMID: 7909431

    		Biochem J. 1994 Apr 1;299 ( Pt 1):309-15.
			Phosphorylation by protein kinase C and
			cyclic AMP-dependent protein kinase of synthetic
			peptides derived from the linker region
			of human P-glycoprotein.

			Chambers TC(1), Pohl J, Glass DB, Kuo JF.

			Author Information
			(1) Department of Pharmacology, Emory University
			School of Medicine, Altanta, GA 30322.

			Specific sites in the linker region of human
			P-glycoprotein phosphorylated by protein
			kinase C (PKC) were identified by means
			of a synthetic peptide substrate, PG-2,
			corresponding to residues 656-689 from this
			region of the molecule. As PG-2 has several
			sequences of the type recognized by the
			cyclic AMP-dependent protein kinase (PKA),
			PG-2 was also tested as a substrate for
			PKA. PG-2 was phosphorylated by purified
			PKC in a Ca2+/phospholipid-dependent manner,
			with a Km of 1.3 microM, and to a maximum
			stoichiometry of 2.9 +/- 0.1 mol of phosphate/mol
			of peptide. Sequence analysis of tryptic
			fragments of PG-2 phosphorylated by PKC
			identified Ser-661, Ser-667 and Ser-671
			as the three sites of phosphorylation. PG-2
			was also found to be phosphorylated by purified
			PKA in a cyclic AMP-dependent manner, with
			a Km of 21 microM, and to a maximum stoichiometry
			of 2.6 +/- 0.2 mol of phosphate/mol of peptide.
			Ser-667, Ser-671 and Ser-683 were phosphorylated
			by PKA. Truncated peptides of PG-2 were
			utilized to confirm that Ser-661 was PKC-specific
			and Ser-683 was PKA-specific. Further studies
			showed that PG-2 acted as a competitive
			substrate for the P-glycoprotein kinase
			present in membranes from multidrug-resistant
			human KB cells. The membrane kinase phosphorylated
			PG-2 mainly on Ser-661, Ser-667 and Ser-671.
			These results show that human P-glycoprotein
			can be phosphorylated by at least two protein
			kinases, stimulated by different second-messenger
			systems, which exhibit both overlapping
			and unique specificities for phosphorylation
			of multiple sites in the linker region of
			the molecule.

			PMCID: PMC1138056 PMID: 7909431 [Indexed
			for MEDLINE]

     			                         Tweet       Print