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@ARTICLE{Chan:637345,
author = {Chan, Hsin-Ju and Krichel, Boris and Bandura, Liam J. and
Chapman, Emily A. and Rogers, Holden T. and Fischer, Matthew
S. and Roberts, David S. and Gao, Zhan and Wang, Man-Di and
Wu, Jingshing and Uetrecht, Charlotte and Jin, Song and Ge,
Ying},
title = {{S}tructural {H}eterogeneity of {P}roteoform-{L}igand
{C}omplexes in {A}denosine {M}onophosphate-{A}ctivated
{P}rotein {K}inase {U}ncovered by {I}ntegrated {T}op-{D}own
{M}ass {S}pectrometry},
journal = {Journal of the American Chemical Society},
volume = {147},
number = {34},
issn = {0002-7863},
address = {Washington, DC},
publisher = {ACS Publications},
reportid = {PUBDB-2025-03837},
pages = {30809 - 30819},
year = {2025},
note = {Waiting for fulltext},
abstract = {Adenosine monophosphate-activated protein kinase (AMPK) is
a heterotrimeric complex (αβγ) that serves as a master
regulator of cellular metabolism, making it a prominent drug
target for various diseases. Post-translational
modifications (PTMs) and ligand binding significantly affect
the activity and function of AMPK. However, the dynamic
interplay of PTMs, noncovalent interactions, and
higher-order structures of the kinase complex remains poorly
understood. Herein, we report for the first time the
structural heterogeneity of the AMPK complex arising from
ligand binding and proteoforms─protein products derived
from PTMs, alternative splicing, and genetic
variants─using integrated native and denatured top-down
mass spectrometry (TDMS). The fully intact AMPK
heterotrimeric complex exhibits heterogeneity due to
phosphorylation and multiple adenosine monophosphate (AMP)
binding states. Native TDMS delineates the subunit
composition, AMP binding stoichiometry, and higher-order
structure of AMPK complex, while denatured TDMS
comprehensively characterizes the proteoforms and localizes
the phosphorylation site. Notably, by integrating native
TDMS and AlphaFold, we elucidate a flexibly connected
regulatory region of AMPK β subunit that was previously
unresolvable with traditional structural biology tools. Our
findings offer new perspectives on protein kinase regulation
and establish a versatile framework for comprehensive
characterization of proteoform-ligand complexes.},
cin = {CSSB-LIV/DESY-CU},
ddc = {540},
cid = {$I:(DE-H253)CSSB-LIV_DESY-CU-20220525$},
pnm = {633 - Life Sciences – Building Blocks of Life: Structure
and Function (POF4-633) / Top-AMPK - Putting the top down on
AMPK protein complex (101068151) / SPOCkS MS - Sampling
Protein cOmplex Conformational Space with native top down
Mass Spectrometry (759661)},
pid = {G:(DE-HGF)POF4-633 / G:(EU-Grant)101068151 /
G:(EU-Grant)759661},
experiment = {EXP:(DE-MLZ)NOSPEC-20140101},
typ = {PUB:(DE-HGF)16},
doi = {10.1021/jacs.5c06950},
url = {https://bib-pubdb1.desy.de/record/637345},
}