PHOSPHORYLATION OF SERINE-2843 IN RYANODINE RECEPTOR-CALCIUM RELEASE CHANNEL OF SKELETAL-MUSCLE BY CAMP-DEPENDENT, CGMP-DEPENDENT AND CAM-DEPENDENT PROTEIN-KINASE

被引:117
作者
SUKO, J [1 ]
MAURERFOGY, I [1 ]
PLANK, B [1 ]
BERTEL, O [1 ]
WYSKOVSKY, W [1 ]
HOHENEGGER, M [1 ]
HELLMANN, G [1 ]
机构
[1] ERNST BOEHRINGER INST ARZNEIMITTELFORSCH,VIENNA,AUSTRIA
关键词
SKELETAL MUSCLE; PROTEIN KINASE; CALCIUM CHANNEL; SERINE PHOSPHORYLATION; PHOSPHORYLATION SITE; SARCOPLASMIC RETICULUM;
D O I
10.1016/0167-4889(93)90023-I
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
The aim of the present study was to determine the phosphorylation of the purified ryanodine receptor-calcium release channel (RyR) of rabbit skeletal muscle sarcoplasmic reticulum by the cAMP-dependent protein kinase (PK-A), cGMP-dependent protein kinase (PK-G) and Ca2+-, CaM-dependent protein kinase (PK-CaM) and the localization of phosphorylation sites. Phosphorylation was highest with PK-A (about 0.9 mol phosphate/mol receptor subunit), between one-half to two-thirds with PK-G and between one-third and more than two-thirds with PK-CaM. Phosphoamino acid analysis revealed solely labeled phosphoserine with PK-A and PK-G and phosphoserine and phosphothreonine with PK-CaM. Reverse-phase high-performance liquid chromatography (HPLC) of cyanogen bromide/trypsin digests of the phosphorylated RyR (purified by gel permeation HPLC) and two-dimensional peptide maps revealed one major phosphopeptide by PK-A and PK-G phosphorylation and several labeled peaks by PK-CaM phosphorylation. Automated Edman sequence analysis of the major phosphopeptide obtained from PK-A and PK-G phosphorylation and one phosphopeptide obtained from PK-CaM phosphorylation yielded the sequence KISQTAQTYDPR (residues 2841-2852) with serine 2843 as phosphorylation site (corresponding to the consensus sequence RKIS), demonstrating that all three protein kinases phosphorylate the same serine residue in the center of the receptor subunit, a region proposed to contain the modulator binding sites of the calcium release channel.
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收藏
页码:193 / 206
页数:14
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