Drug discovery

Protein Production and Characterisation (PPC) assists researchers with the expression, purification and analysis of their proteins of interest. We facilitate the production and characterisation of proteins, including those that may constitute major drug targets. Services include: 

• undertaking experiments to produce and characterise target proteins in state-of-the-art laboratories
• providing expert advice to users and training personnel (including students and staff). 

We have the expertise to express your protein in bacterial, insect, or mammalian cell host systems. We have access to highly specialised protein purification equipment and chromatography columns, and can tailor protocols to your target of interest.

Download Protein Production and Characterisation (pdf, 906 KB) for more information about our services and instruments. 

We have the equipment to undertake biochemical and biophysical characterisation of samples. Staff can collect measurements, or train users to collect data. Our instruments are described below or download Protein Production and Characterisation (pdf, 906 KB) for more information.

Prometheus Panta

Prometheus Panta is a protein stability screening platform. It combines four different measurement modes: nano differential scanning fluorimetry (nanoDSF), static light scattering (SLS), dynamic light scattering (DLS) and back reflection. This allows thermal unfolding, particle sizing, and aggregation measurements to be done simultaneously. The curated design and quality of the Prometheus Panta make it the current go-to instrument for screening protein stability.

UNCLE

UNCLE is a protein stability screening platform. It combines three different measurement modes: differential scanning fluorimetry (DSF, with or without a fluorophore like SYPRO Orange), static light scattering (SLS), and dynamic light scattering (DLS). This allows thermal unfolding, particle sizing, and aggregation measurements to be done simultaneously, which makes UNCLE one of the most popular instruments for screening protein stability.

Refeyn Two MP Mass Photometer

The Refeyn Two MP Mass Photometer is a relatively new method for molecular analysis. It measures the mass of single molecules in solution using light. It analyses individual molecules or complexes in their native state and label free. It has high sensitivity, a fast acquisition time, and low sample consumption (typically < 100 nM). It can measure proteins, antibodies, nucleic acids, lipids, and small viruses, with a detection range of 30 kDa – 5 MDa.

Circular Dichroism (CD)

CD can be used to assess protein secondary structure, as well as for protein stability studies. It has low sample consumption and a high degree of sensitivity. 

SEC-MALLS

We have access to and experience with running Multi-angle laser light scattering (MALLS) coupled to Size-Exclusion-Chromatography (SEC). An advanced and conformation-independent way of measuring the molar mass and size of species as they elute from a chromatography matrix. 

Protein X-ray crystallography is one of the most powerful methods to visualise the molecular structure of a protein at atomic resolution. The structure, in turn, provides a better understanding of a protein’s functionally important sites and its mechanism of action. This facilitates the development of efficient and specifically targeted drugs.

Sydney Analytical provides a platform for users to crystallise their proteins using state-of-the-art instruments. Our expert staff provide guidance and can assist researchers, especially those new to the field of X-ray crystallography, in determining the three-dimensional structure of their target protein.

Services offered include:

• crystallisation and structure solution (including training)
• data collection at the Australian Synchrotron*
  
• structure determination and refinement.

*Synchrotron X-rays are much more intense than in-house sources, allowing much smaller crystals to be measured much more quickly. Our partners at ANSTO operate three protein crystallography beamlines at the Australian Synchrotron (MX1 and MX2, with MX3 due to open shortly) tuned for different types of problems. We have access to the Australian Synchrotron X-ray facility through the Collaborative Access Program (CAP) which allows for the screening and data collection for crystals.

Download Macromolecular Crystallography (pdf, 417 KB) for more information about available Protein Crystallography equipment and expertise. 

We have equipment to undertake biophysical analysis of a variety of interaction types. Staff can collect measurements, or train users to collect data. Our instruments are described below or download Macromolecular Interactions Analysis (pdf, 240 KB) for more information. 

Blitz (BiLayer Interferometry)

BiLayer interferometry is an optical analytical technique that assesses the interference pattern of white light reflected from two surfaces: a layer of immobilised protein on the biosensor tip, and an internal reference layer. Any change in the number of molecules bound to the biosensor tip causes a shift in the interference pattern that can be measured in real time to determine affinity and kinetic measurements.

Good for measuring: Protein:Protein, Protein:peptide, Protein:Nucleic Acids

Biacore T200 (Surface Plasmon Resonance)

Surface plasmon resonance (SPR) is an optical technique that can be used to measure interactions in real-time. SPR occurs when polarised light strikes an electrically conducting surface at the interface between two media. This generates electron charge density waves called plasmons, reducing the intensity of reflected light at a specific angle known as the resonance angle, in proportion to the mass on a sensor surface.

A typical experiment involves one binding partner (ligand) immobilised to the surface of an SPR sensor chip, either directly or via an affinity tag such as biotin.  The other partner (analyte) is then flown over the surface in increasing concentrations. If an interaction occurs, the change in mass on the sensor surface affects the reflected light, which is detected and plotted as an output sensorgram. This allows calculation of the affinity and kinetics of the interaction.

Good for measuring: Protein:Protein, Protein:peptide, Protein:Nucleic Acids, Protein:Small Molecules

PEAQ MicroCal ITC (ITC)

Isothermal Titration Calorimetry (ITC) measures the binding affinity between any two molecules that either release or absorb heat upon a binding interaction occurring. 

The microcalorimeter instrument measures the heat difference between a sample cell and a reference cell (containing water or buffer) that occurs upon titration of increasing amount of a binding partner. ITC can be used to measure a variety of thermodynamic properties of biomolecular interactions, including affinity (KD), stoichiometry(n), as well as enthalpy (DH) and entropy (DS).

Good for measuring: Protein:Protein, Protein:peptide, Protein:Nucleic Acids, Protein:Small Molecules

Fragment-based drug discovery (FBDD) is a strategy that is growing rapidly in popularity. Rather than rely on large libraries of complex molecules, FBDD involves a small, carefully curated library of molecules that are about half the size of typical drugs. A drug target is screened against the library and ‘hits’ can then be chemically expanded to create lead compounds that bind with high affinity. The approach can be used to target enzymes as well as less traditional targets such as protein-protein or protein-nucleic acid interactions, and even membrane proteins.

Sydney Analytical use a fragment library curated by the medicinal chemists at the Monash Fragment Platform. Our primary screening methods are Nuclear Magnetic Resonance (NMR) and Surface Plasmon Resonance (SPR). Please download Fragment Based Drug Discovery (pdf, 154 KB) for more information about our FBDD workflow, as well as our equipment and expertise.

Our specialised Cyclic Peptide Screening service introduces you to the remarkable potential of small cyclic peptides in the realm of drug discovery. These compact yet powerful peptides are designed to adeptly disrupt interactions while embodying drug-like properties.

The efficiency of our Cyclic Peptide Display Screening (CPDS) service, a cutting-edge platform that seamlessly combines rapid parallel synthesis and screening. This innovative approach allows for the exploration of trillions of molecules against a wide array of expressible protein targets. The outcome? A curated collection of diverse ligands boasting exceptional affinity (typically low nM) and selectivity.

In the landscape of drug discovery, our service shines a spotlight on cyclic peptides, offering you a personalised and scientific approach. Uncover the potential of these molecular gems as we guide you through the intricacies of CPDS, paving the way for groundbreaking opportunities in the pursuit of tailored medications.

For more information, please download Cyclic Peptide Display Screening (pdf, 517 KB)

In vitro cell cultures are crucial tools for modelling diseases. 3D cell cultures more accurately model human physiology and provide better insights for oncology, drug discovery, and personalised medicine applications. We can assist with designing and developing automated 3D culture platforms for high-throughput drug discovery screening, personalised medicine screening, advanced therapeutics, and investigative biological studies.

Services

· Automation cell culture for human pluripotent stem cell (iPSC) cell lines such as cardiomyocytes, astrocytes, hepatocytes, retinal ganglion cells, kidney cells, intestinal cells, and airway cells) in multiple 3D structures (spheroids, organoids, and organ-on-chip).

· Automation cell-based high-content/high-throughput drug screening assays using either the spinning confocal imaging system or multi-mode microplate readers.

· Quantitative determination of cell-based permeability assay (for example Caco-2) using mass spectrometry analysis for drug screening.

Equipment

We have a dedicated lab space and instrumentation to assist with cell culture. This includes:

· Biosafety and cyBioSafety Class II and Cytotoxic cabinets 

· Bio-Rad micropulser and neon electroporators 

· Countess cell counter  

· Cytation

· Perkin Elmer Janus Liquid Handling Robot

RASTRUM™: A 3D cell bioprinter that utilises different/multiple cell types (primary cells and iPSCs) in a tunable 3D matrix (single or multiple models) that closely mimics the complexity of physiological and pathological in vivo microenvironments for next-generation drug discovery and phenotypic research.

Multi-mode microplate readers

We have multi-mode microplate readers that come with built-in absorbance (ABS), luminescence (LUM), fluorescence intensity, fluorescence polarization (FP), and time-resolved fluorescence (TRF) modes. Compatible with 6- to 384-well microplates.

Detection technologies performance:

· Absorbance (230 – 1000 nm)

· Luminescence (300 – 850 nm)

· Fluorescence intensity Ex (250 – 830 nm) / Em (270 – 850 nm); FI filter 440/480 (thioflavin)

· Fluorescence polarization Ex (400 – 750 nm) / Em (400 – 750 nm); FP filters 485/520/520 nm (fluorescein), 540/590/590 nm (TAMRA/CY3), 590/675/675 nm (Cy5)

· Time-resolved fluorescence Ex filter 350 nm / Em (450 – 750 nm) or filter 490 nm (Terbium), 616 nm (Europium)

· HTRF filter 337/665/620 nm

· AlphaScreen filter 680/570 nm; AlphaLISA filter 680/615 nm

The following is a complete list of all the equipment from the drug discovery node that is available to researchers. Contact us at sydney.analytical@sydney.edu.au for more information about any of the equipment listed below.

• Incushakers and incubators for bacterial, insect and mammalian culture
• Avestin Emulsiflex C3 homogeniser for cell lysis
• AKTA Start FPLC Purification system
• AKTA FPLC purification systems
• Beckman Allegra Swing Bucket Centrifuge
• Biocomp Gradient station 
• Beckman Optima L-80XP ultracentrifuge 
• BioRad Trans-Blot Turbo transfer system 
• Apparatus for running DNA agarose gels
• Apparatus for running SDS-PAGE gels
• Bio-rad T100 thermal cycler 
• RotorGeneQ (qPCR machine)
• Mic qPCR machine
• Illumina iSeq Nucleic Acid Sequencer
• Unchained Labs UNCLE for DLS/DSF
• NanoTemper Prometheus Panta for DLS and/or nanoDSF
• Jasco Circular Dichroism (CD) spectropolarimeter
• Refeyn Two MP Mass photometer
• Blitz (Bi-layer interferometry)
• Biacore T200 (Surface Plasmon Resonance)
• MicroCal PEAQ Isothermal Titration Calorimeter (ITC)
• Protein Crystallisation equipment:
  • Includes an NT8Ⓡ (Formulatrix), and access to a MosquitoⓇ LV (sptlabtech), Freedom Evo liquid handler (Tecan), Stereo Microscopes (Leica) and Incubators at various temperatures (Memmert)
• PC1 and PC2 Lab space 
• BioSafety Class II and Cytotoxic cabinets 
• Bio-Rad micropulser and neon electroporators 
• Countess cell counter  
• IncuCyte Zoom 
• Perkin Elmer Janus Liquid Handling Robot
• Rastrum 3D Cell Culture Platform
• Microplate readers with built-in absorbance (ABS), luminescence (LUM), fluorescence intensity, fluorescence polarization (FP), and time-resolved fluorescence (TRF) modes. Compatible with 6- to 384-well microplates