Chemistry Reference
In-Depth Information
Besides backbone dynamics for monomeric proteins, ensemble minimization may
be better suited for studying relatively slower and larger scale protein domain dyna-
mics and structure. Liu et al. [
78
] implemented ensemble minimization to charac-
terize the dynamic structure of a membrane-anchored ADP ribosylation factor (Arf).
The yeast Arf is composed of flexibly joined C-terminal domain and the short
N-terminal helix that interacts with the membrane. The Arf protein anchored on
DMPC/DHPC bicelles were aligned in a negatively charged polyacrylamide gel,
and the resulting alignment order parameters
D
a
and
R
for C- and N-terminal domains
are 8.6 Hz and 0.29, and 4.7 Hz and 0.61, respectively. The differences in alignment
order indicate significant interdomain motions. A three-member ensemble minimiza-
tion significantly increased the linear correlation coefficient between measured and
back-calculated RDC (Fig.
6
) and PRE data on the N-terminal helix. Such dynamics
could be crucial for Arf to function as a regulator of effectors GTP/GDP.
4.4 Structure of a Ligand in a Bound State
The conformational ensemble described previously represents a continuous distri-
bution of conformers and no significant protein fold or high-order structural change
within the ensemble members was taken into account. Other systems may also
be composed of discrete conformers in equilibrium; for instance, large structural
differences could exist between free and bound states of ligand molecules in the
presence of their receptors. Generally the NMR signals from the ligands in this type
of equilibrium are dominated by the ligands in the free states. RDCs can play a role in
solving ligand structure in the bound formnot generally accessible by other techniques.
Photo-activated retina rhodopsin, a G-protein coupled receptor (GPCR), is
a major component in purple membranes (PM). Due to the high structural content
of helices in GPCR, GPCR rich PM disks align in the presence of a magnetic field
Fig. 6 Ensemble structural fitting to RDCs. Agreement between experimental and back-
calculated RDCs for a one-state ensemble (
left
) and a three-state ensemble (
right
). Data include
NH, NC
0
, and phenyl CH (the latter two are normalized to NH). RDCs are collected in positive and
negative gels. (Reprinted with permission from [
78
])
