Download G-protein coupled receptor over-expression in

Understanding structure-activity relationships in G-protein coupled receptors (GPCRs)
Dr Roslyn Bill and Dr David Poyner
http://www.aston.ac.uk/lhs/staff/A-Zindex/billrm.jsp
http://www.aston.ac.uk/lhs/staff/A-Zindex/poynerdr.jsp
GPCRs are the single largest protein family in the mammalian genome, and the largest class of drug
targets. Unfortunately, they are only available in minute quantities in the cell (typically less than 0.1% of
the protein complement). It is therefore recognised by the scientific community that the only way to
overcome the bottleneck in gaining a complete understanding of GPCRs and thus to develop new drugs is
to produce sufficient quantities of this key class of proteins for structural and functional analysis.
Currently, the only available high-resolution structure of a GPCR is that of rhodopsin, the chief
vertebrate photoreceptor, which unusually for a membrane protein is present in large quantities in the cell.
It is clear that this structural information cannot necessarily be used to model other GPCRs. Progress in
this field therefore relies on the recombinant production of large quantities of pure, functional membrane
protein for biochemical and structural studies. The proposed project therefore unites the protein
expression expertise of Roslyn Bill with the pharmacological expertise of David Poyner to investigate
and develop eukaryotic expression systems to produce both full length and soluble forms of a unique
GPCR, the CL-receptor, and its accessory protein, RAMP1.
Roslyn Bill is known for her work on understanding the molecular basis of successful membrane
protein overexpression in the eukaryotic hosts, Saccharomyces cerevisiae and Pichia pastoris. Her group
has successfully produced a range of membrane proteins for crystallisation trials and subsequent structural
analysis. The main theme of her work is in the development of novel expression hosts, for which she has
submitted patent applications. Bill’s group will use these and other yeasts to produce CL and RAMP1 for
structural and functional analysis.
David Poyner has been working on the molecular pharmacology of CL/RAMP1, which is implicated
in neurogenic inflammation, migraine and various cardiovascular disorders, for 15 years. The mode of
action of RAMP1 is unknown but it and related RAMPs have now been shown to associate with around
10 GPCRs. Poyner’s group has successfully modelled both CL and RAMP1 and thus can suggest how
they associate to produce functional receptors.
The resulting proteins will be investigated structurally using NMR by Dr. B. Birdsall at the National
Institute for Medical Research, and by X-ray crystallography by Dr Richard Neutze at Chalmers
University, Gothenburg, Sweden. The project will be the catalyst for subsequent joint applications from
Drs Bill and Poyner to the BBSRC and the Wellcome Trust, to investigate the structural basis of other
GPCRs and their interactions with RAMPs, thereby laying the foundation for a deeper understanding of
their pharmacology and facilitating novel drug design;