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@ARTICLE{Levorin:1042312,
author = {Levorin, Leonardo and Becker, Nina and Uluca-Yazgi, Boran
and Gardon, Luis and Kraus, Mirko and Sevenich, Marc and
Apostolidis, Athina and Schmitz, Kai and Rüter, Neomi and
Apanasenko, Irina and Willbold, Dieter and Hoyer, Wolfgang
and Neudecker, Philipp and Gremer, Lothar and Heise,
Henrike},
title = {{I}soleucine {S}ide {C}hains as {R}eporters of
{C}onformational {F}reedom in {P}rotein {F}olding {S}tudied
by {DNP}-{E}nhanced {NMR}},
journal = {Journal of the American Chemical Society},
volume = {147},
number = {18},
issn = {0002-7863},
address = {Washington, DC},
publisher = {ACS Publications},
reportid = {FZJ-2025-02519},
pages = {15867 - 15879},
year = {2025},
abstract = {Conformations of protein side chains are closely linked to
protein function. DNP-enhanced solid-state NMR (ssNMR),
which operates at cryogenic temperatures (<110 K), can be
used to freeze-trap protein conformations, including the
side chains. In the present study, we employed
two-dimensional DNP-enhanced ssNMR to get detailed insights
into backbone and side chain conformations of isoleucine. We
used different amino acid selectively labeled model proteins
for intrinsically disordered proteins (IDPs), denatured and
well-folded proteins, and amyloid fibrils. 13C chemical
shifts are closely correlated with secondary structure
elements and χ1 and χ2 angles in isoleucine side chains.
Thus, line shape analysis by integration of representative
peak areas in 2D spectra provides an accurate overview of
the distribution of backbone and side chain conformations.
For the well-folded proteins GABARAP and bovine PI3-kinase
(PI3K) SH3 domain, most Ile chemical shifts in frozen
solution are well resolved and similar to those observed in
solution. However, line widths of individual Ile residues
are directly linked to residual mobility, and line
broadening or even signal splitting appears for those Ile
residues, which are not part of well-defined secondary
structure elements. For unfolded PI3K SH3 and the IDP
α-synuclein (α-syn), all Ile side chains have full
conformational freedom, and as a consequence, inhomogeneous
line broadening dominates the cryogenic spectra. Moreover,
we demonstrate that conformational ensembles of proteins
strongly depend on solvent and buffer conditions. This
allowed different unfolded structures for chemical and
acidic pH denaturation of the PI3K SH3 domain to be
distinguished. In amyloid fibrils of α-syn and PI3K SH3,
chemical shifts typical for β-strand like secondary
structure dominate the spectra, whereas Ile residues
belonging to the fuzzy coat still add the IDP-type line
shapes. Hence, DNP-enhanced ssNMR is a useful tool for
investigating side chain facilitated protein functions and
interactions.},
cin = {IBI-7},
ddc = {540},
cid = {I:(DE-Juel1)IBI-7-20200312},
pnm = {5244 - Information Processing in Neuronal Networks
(POF4-524)},
pid = {G:(DE-HGF)POF4-5244},
typ = {PUB:(DE-HGF)16},
pubmed = {40285725},
UT = {WOS:001477553300001},
doi = {10.1021/jacs.5c04159},
url = {https://juser.fz-juelich.de/record/1042312},
}