Chemistry Reference
In-Depth Information
advantages: (1) it is composed of a low number of small, isolated spin systems, (2)
transfer of magnetization into the side chain is thus eliminated and spectral quality
enhanced, and (3) the number of inter-residue cross-peaks is significantly increased
which is important both for assignment and structure calculation [
49
]. This labeling
scheme or similar ones may be applicable to other large membrane proteins, e.g.,
the 7-transmembrane family proteins.
Another great achievement by SSNMR is the structure determination of the
HET-s (218-289) fibril by the Meier group [
42
], as shown in Fig.
5
. Total 90
13
C-
13
C and 44
1
H-
1
H distance restraints obtained by the CHHC, NHHC and
PDSD experiments, and 74 angle restraints obtained by TALOS [
193
], were used
for structure calculations. The extraordinarily high order in the HET-s prion fibrils
can be explained by the well-organized structure obtained by SSNMR.
3.3 Ligand Conformation and Binding
The molecular mechanisms of membrane protein activation are at the center of
interest in the study of cellular responses to biogenic stimulus and drugs. For
example, GPCRs are activated by a wide range of stimuli, including hormones,
neurotransmitters, ions, odorants, and photons of light [
128
]. Knowledge of the
three-dimensional structures of several GPCRs, such as rhodopsin,
b
2
AR,
A
2a
R, CXCR4, D
3
R, and H
1
R[
129
-
141
] have been resolved by X-ray crystallog-
raphy in either an inactive state or agonists/antagonists bound form at high resolu-
tion which open up new possibilities for investigating GPCRs of human therapeutic
significance and for rational drug design. However, despite the availability of those
crystal structures, a comprehensive understanding of the mechanism of structure
activation is still a challenge due to the lack of high resolution structure at the
activated state. For example, activation of rhodopsin has still been challenging
because of the lack of high resolution structure at atomic level for the activated
Meta II state [
6
]. On the other hand, SSNMR can offer direct measurements at
atomic resolution to study protein activation caused by the conformational changes
and the ligand binding interactions [
65
,
120
].
The conformation of retinal chromophore and protein activation upon the 11-
cis
to all-
trans
isomerization of the retinal have been studied extensively by MAS
SSNMR through the chemical shift measurements, distance measurements, corre-
lation experiments, and
2
HNMR[
12
-
18
,
20
,
22
,
23
,
25
,
95
,
103
,
104
,
142
-
144
].
Complete assignment of the retinal carbons in ground state and partially in the
Batho-, Meta I, and Meta II intermediates have been achieved [
14
,
17
,
145
-
151
].
These valuable data allow us insight into the conformational changes of the retinal
protonated Schiff base (PSB) complex and the related transmembrane helices
through the transition from the ground state to the Batho, Meta I, and Meta II
intermediates in the binding pocket, as shown in Fig.
6
.
Large down-field or up-field changes have been observed at the C16, C17 of the
b
1
AR,
b
-ionone ring, and on the retinal polyene chain around C9-C10-C11-C12 and
