Protein structural information provides a solid foundation for the
discovery and development of drugs that will be highly selective and effective
against chosen targets. As a leading provider of structural biology services,
deCODE biostructures utilizes proprietary technology and methods to impart
innovation to every client project. Our capabilities span the entire
gene-to-structure landscape including:
Computer-Aided Design of Expression-Optimized Synthetic
Genes Engineering of protein constructs for high-level production
and efficient crystallogenesis can be achieved for client projects using Gene
Composer™, a powerful database software package for synthetic gene design
developed and coded by scientists at deCODE biostructures.
Whole gene
synthesis is rapidly becoming a powerful technology for improving protein
expression. The design of optimal nucleic acid sequences for specific expression
systems can be extremely complicated due to the many factors that must be
considered simultaneously. Specifically, consideration should be made for
appropriate codon usage frequency that will not result in mRNA secondary
structures that may inhibit translation. Additionally there are many subtle
features that can impact the quality and yield of protein including unusual
sequence repeats, out-of-frame stop codons, and cryptic regulatory sequences
(e.g. Shine Dalgarno, splice sites, RNase cleavage sites, etc.).
Using
Gene Composer™, deCODE biostructures can rapidly create optimized nucleic acid
sequences for any expression system. This database tool can also be used to
design unique and stable proteins by creating multiple sequence alignments and
distilling protein structure information.
Parallel Protein Purification and Analysis The
protein production laboratories of deCODE biostructures are equipped with
state-of-the-art fermentation systems for production of recombinant protein in
E. coli, yeast, baculovirus-insect cells, and mammalian cell expression systems.
Laboratories are also equipped with Protein Maker™ automated systems which can
purify 24 proteins simultaneously from crude cell extracts. Protein Maker™ is
another proprietary technology developed at deCODE biostructures to address
bottlenecks in protein purification. This system is used for both scouting and
preparative work and also allows sequential multi-column preparative
purifications on up to 8 proteins simultaneously. This tool gives deCODE
biostructures a distinct advantage in high quality protein purification for
crystallography. deCODE biostructures maintains numerous traditional protein
chromatography workstations and HPLC systems for large-scale purification
projects. All purified proteins are rigorously tested for quality assurance to
assess protein purity, aggregation state, and activity.
The protein
quality assurance laboratory maintains sophisticated analytical instruments for
enzyme assays, dynamic light scattering, mass spectrometry, UV
spectrophotometry, and amino acid sequencing.
High-Throughput Crystallization Screening High-throughput crystallization is performed in a core lab equipped with
sophisticated liquid handling devices including Matrix Maker™ and Drop Maker™
which were developed by deCODE biostructures to quickly and accurately prepare
crystallization screening kits and set-up crystallization experiments in plate
format. Specifically, our scientists developed Clover 384™ an innovative
384-well plate that facilitates a combinatorial approach for efficient
identification of crystal growth conditions including those that depend on the
presence of ligands (co-crystallization conditions), as well as others that
produce Apo-crystal forms for soaking with ligands to provide protein-ligand
co-crystal structures. Crystallization methodologies include vapour diffusion,
batch crystallization, free interface diffusion and several other novel methods
to increase crystal hit rate and improve crystal quality. Thousands of
crystallization experiments are monitored every day in laboratories equipped
with CrysTEL™ plate hotels and Crystal Monitor Workstations™ with DETECT-X™
automated crystal detection systems. All of these modern automation technologies
were developed at deCODE biostructures and are available commercially. These
unique tools and methods are integrated to create a high-output engine for
crystal production.
X-ray Data Collection deCODE biostructures
maintains state-of-the-art X-ray diffraction data collection facilities in house
that allow the collection of several complete X-ray diffraction data sets on
typical crystals each day. deCODE also maintains numerous proprietary
synchrotron beamline agreements that allow our scientists to collect X-ray
diffraction data at national laboratories with synchrotron facilities equipped
with protein crystal X-ray diffraction data collection instruments.
Protein-Ligand Co-Crystal Structure Determination The timely availability of X-ray crystal structures that reveal ligand
binding modes with their protein targets can provide a wealth of information for
medicinal chemistry. deCODE biostructures is an experienced and trusted provider
of structural biology information to support the medicinal chemistry programs of
clients. deCODE biostructures has assembled talented researchers, automation,
software, and methods that are specifically suited to delivery of co-crystal
structures for numerous target classes of proteins including kinases,
phosphatases, proteases, polymerases, topoisomerases and others.
Crystal Ready™ Targets Clients can now take
advantage of the successful experience of deCODE biostructures in resolving
ligand-bound co-structures for a growing number of Crystal Ready™ targets
(www.crystalready.com). Crystal Ready™ targets are active proteins that are
available in formulated soluble form for co-crystallization with ligands, or in
stabilized crystalline form for soaking with ligands. Our scientists have
validated Crystal Ready™ targets for successful co-crystal structure
determination by either co-crystallization or soaking with a variety of control
compounds with affinities that span several orders of magnitude.
Difficult Targets & Membrane Protein Research Integral membrane proteins represent approximately half of all major drug
targets. These include G-Protein Coupled Receptors (GPCRs) and Ion Channels
which are notoriously difficult to produce, purify, and crystallize. deCODE
biostructures is one of a limited number of companies that are making major
technology investments directed at crystallographic structure determinations of
ligand-bound GPCRs and Ion Channels. Specifically, deCODE biostructures has
developed unique and proprietary methods for the crystallization of integral
membrane proteins in their lipid bilayer environment. This method for small
volume Lipidic Cubic Phase crystallization allows our scientists to set up
thousands of crystallization experiments using less than 1 mg of protein and can
be accomplished at 10 nL volumes, a unique ability in the industry. deCODE
biostructures also developed the DETECT-X™ microscope system, a powerful
automated imaging tool that aids in detecting non-colored, micrometer size
crystals. Additionally, deCODE biostructures’ synthetic gene methods and special
insect cell lines enable greatly improved production of active recombinant human
GPCRs. This scouting and optimization approach has been validated for both class
A and B GPCRs. As an innovator in this rapidly developing discipline, deCODE
biostructures has made a long term commitment to membrane protein
crystallography and research.
Advancements in this discipline include:
Heterologous expression in recombinant baculovirus-insect cell culture and
mammalian cell culture, as well as E.coli and yeast.
Membrane protein stabilization (Activity assays, detergents, scouting)
Co-expression of GPCRs with G-proteins (vector sets available)
Expression with alternate signal peptides
Rigorous optimization approach at the stages of expression, purification and
crystallization
Insect cell line Sf9-XL (developed in house) and also available Sf-9, Hi-5,
Sf-21, and proprietary culture medium
Multiple 10 liter wave bag fermentation
Membrane fractionation by gradient centrifugation
Modification of glycosylation pattern (enzymatic, culture conditions)
Crystallization in the Lipidic Cubic Phase micro-method
