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No mean feat
September 2013
EDIT CONNECT
SHARING OPTIONS:
MELBOURN, United Kingdom—For the first time, researchers
have been able to successfully report the structure of a Class B G-Protein
Coupled Receptor (GPCR) using lipidic cubic phase (LCP) protein
crystallization. The work performed at Heptares Therapeutics Ltd.,
a drug
discovery and development company, and published recently in Nature, details the structural identification of the stress
receptor, corticotropin-releasing factor receptor 1 (CRF1), which opens up
the
technique to an entirely new group of molecules with important roles in the
development of diseases as diverse as diabetes, osteoporosis,
depression and
anxiety. It is hoped that the Class B GPCRs will become valuable drug targets.
The
mosquito LCP, a solution for LCP crystallography
designed by TTP
Labtech, was a significant technical driving force enabling the
research.
"TTP Labtech's mosquito
LCP overcomes the common problems
encountered with accurately dispensing the highly viscous LCP mixture used in
membrane protein crystallization,"
notes Joby Jenkins, mosquito product manager
and global director of automation at TTP Labtech. "It allows you to fully
automate LCP set-ups accurately
and repeatedly, dispensing LCP volumes as low
as 25 nL, while automated calibration of syringe and pipette positioning
ensures precise drop-on-drop
placement for easy automated imaging."
Basically, he says, the mosquito operates on the principle
of positive displacement. The viscous LCP lipid is heated with a protein in the
absence of air while in contact with a plunger, which then deposits
the mixture
on a glass membrane. An auto calibration feature ensures the accurate placement
of each drop.
GPCRs are a family of proteins that play an essential part
in cell signaling and are thus important drug targets for
modulating cell
function and influencing disease outcome. The Class B subset of GPCRs includes
many peptide hormone receptors relevant for treating
disease, such as glucagon,
glucagon-like peptide, calcitonin and parathyroid peptide hormone. However,
until the recent work carried out by Heptares,
it had proven almost impossible
to provide structural data for this class, severely hampering drug development
efforts across the industry.
The finding that the structure of CRF1, a Class B GPCR,
contains a novel binding pocket for the small-
molecule antagonist, towards the
intracellular side of the receptor and far from the position of other Class A
GPCR ligands, underlines the importance
of the discovery, particularly in terms
of modeling other Class B receptors and potentially unlocking drug design
across the subset.
Generating crystalline structures of key biological
receptors is an important step in drug discovery, as it enables
protein-protein
interactions to be reliably predicted and fuels the rational design of
molecules engineered to influence receptor activity. However,
protein
crystallography, often referred to as the "black art," is frequently a long and
complex process requiring great technical skill, precision
and patience. This
is especially true when working with complex transmembrane structures such as
GPCRs, which require the optimization of specific
experimental and
environmental conditions for each discrete structure.
Dr. Andrew S. Doré, head of
crystallography at Heptares and
co-author of the recent article in Nature,
said, "In any crystallographic project, eliminating variables is
key in
producing a steady stream of crystallization-grade protein for structure
determination. This, of course, also extends to crystallization itself.
We had
to stitch together data from 35 isomorphous crystals to produce a full dataset
for the CRF1 structure solution. Mosquito LCP provided a reliable
platform for
our CRF1 LCP crystallization, deploying reproducible low volume boli at speed,
and ultimately yielding a large sample set of CRF1 crystals
for subsequent
diffraction analysis."
TTP Labtech's mosquito LCP is used by many leading
pharmaceutical and biotechnology companies for protein crystallization
screening and production, playing an important role in the identification of
many protein structures, including important GPCRs. However, this is perhaps
the most significant, high-profile discovery published to date using the
technology.
"We've worked closely with Doré's team at Heptares since
2010—in fact, they were
one of the earliest adopters of the mosquito LCP,"
noted Jenkins. "Heptares has now solved in excess of 35 GPCR structures
to-date—no mean feat when
working with such a complex experimental process." Code: E091320 Back |
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