Amide H/2H exchange reveals communication between the cAMP and catalytic subunit-binding sites in the RIα subunit of protein kinase A

被引:85
作者
Anand, GS
Hughes, CA
Jones, JM
Taylor, SS
Komives, EA
机构
[1] Univ Calif San Diego, Dept Chem & Biochem, La Jolla, CA 92093 USA
[2] Univ Calif San Diego, Howard Hughes Med Inst, La Jolla, CA 92093 USA
关键词
MALDI-TOF; H/H-2; exchange; protein kinase; signal transduction; cAMP;
D O I
10.1016/S0022-2836(02)00919-1
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 changes in backbone hydrogen/deuterium (H/H-2) exchange in the regulatory subunit (R(I)alpha(94-244)) of cyclic AMP-dependent protein kinase A (PKA) were probed by MALDI-TOF mass spectrometry. The three naturally occurring states of the regulatory subunit were studied: (1) free R(I)alpha(94-244), which likely represents newly synthesized protein, (2) R(I)alpha(94-244) bound to the catalytic (C) subunit, or holoenzyme, and (3) R(I)alpha(94-244) bound to cAMP. Protection from amide exchange upon C-subunit binding was observed for the helical subdomain, including the A-helix and B-helix, pointing to regions adjacent to those shown to be important by mutagenesis. In addition, C-subunit binding caused changes in observed amide exchange in the distal cAMP-binding pocket. Conversely, cAMP binding caused protection in the cAMP-binding pocket and increased exchange in the helical subdomain. These results suggest that the mutually exclusive binding of either cAMP or C-subunit is controlled by binding at one site transmitting long distance changes to the other site. (C) 2002 Elsevier Science Ltd. All rights reserved.
引用
收藏
页码:377 / 386
页数:10
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