Protein
characterization.
We are protein biodesigners
Synthelis offers nearly all available methods, from the simplest to the most sophisticated techniques, to characterize your proteins of interest from both a functional and structural point of view. We use either our own equipment or partner platforms. Depending on your project, we can help you determine the best technique to quantify and qualify your protein, to study its activity or analyze the interactions it has with ligands, or to determine its three-dimensional structure.
Below is a list of the methods we use either as standard (SDS-PAGE and Western Blotting) or as an option:
Qualitative studies
To guarantee the quality of the proteins we have synthesized for you or that you have produced, we offer our qualitative analysis services. These analyses allow us to determine purity levels for proteins of interest, to validate their identity and to study the homogeneity of the solutions obtained.
Certain analyses such as SDS-Page control and Western-blot analysis are included in our protein production offerings to ensure the purity and identity of the product obtained.
Depending on your needs, we can complete this offering with different methods allowing a more in-depth study. We can check the identity and integrity of the protein using mass spectrometry, analyze the oligomerization state by SEC, SEC-MALS or DLS and study the stability of the protein by TSA. We can also check for the presence of endotoxins and verify the sterility of the product.
Methods – Protein/Sample Information
SDS-PAGE (reducing and non-reducing conditions) – Purity
Western blotting – Identity and integrity
Mass Spectrometry – Exact molecular weight and sequence
Size Exclusion Chromatography (SEC) – Oligomeric status and purity
Thermal Shift Assay (TSA) – Protein stability
LAL – Endotoxin level determination
Sterility testing – Absence of viable microorganisms
RP- & SE-HPLC – Identity and purity
nanoDSF – Stability and folding
Quantitative studies
In order to better characterize the proteins produced in our systems, be they cell-free or cellular, we offer different quantification methods to meet your specific needs.
We include, as standard, a UV assay to determine the protein concentration obtained in solution, or for proteins produced in proteoliposomes, a quantification on SDS-PAGE gel.
Depending on your projects, we can also perform other quantifications such as Bradford or Pierce type colorimetric assays and amino acid assays.
Methods – Protein/Sample Information
SDS-PAGE – Specific quantities
UV assay – Total quantity of protein
Colorimetric assay – Total quantity of protein
Amino acid assay – Total quantity
Interaction studies
Interactions between proteins lie at the heart of most cellular processes and are essential for a large number of functions in the living world. These interactions are notably involved in signal transduction, protein complex formation, protein transport, activation or inhibition of metabolic pathways etc.
Synthelis can offer interaction studies of your protein of interest with its ligand. There are many methods to investigate protein-protein interactions with each method having its own advantages and disadvantages. We offer a wide variety of techniques: SPR, BLI, ITC, FRET or MST so that clients can choose the most suitable technique for their project.
We have access to the following instruments for these analyses:
Monolith NT.115 (MST)
Octet® RED96e (BLI)
Biacore T200 (SPR)
PEAQ-ITC (very sensitive and requires small sample volumes)
Methods – Protein/Sample Information
ELISA
Microscale thermophoresis (MST) – Specificity and affinity
BLI – Specificity and affinity
SPR – Specificity, Kd, Kon, Koff
PEAQ-ITC – Binding stoichiometry (n), affinity constant (KD), and enthalpy (ΔH) and entropy (ΔS) changes
Activity studies
We can verify the activity of your proteins through enzymatic activity assays, electrophysiological assays on Orbit Mini (Nanion) and we can also, depending on your needs, develop and perform custom activity assays.
Methods – Protein/sample information
Patch-clamp – Channel activity
Enzyme activity – Functionality
Custom assays – Functionality
Radio ligand binding – Specific activity
Structural studies
Finally, we can offer you several structural analyses, ranging from the evaluation of secondary structures by Circular Dichroism (CD) or Fourier Transform Infrared Spectroscopy (FTIR), to the resolution of three-dimensional structure by cryo-electron microscopy (Cryo-EM), Nuclear Magnetic Resonance (NMR) spectroscopy or by crystallography and X-ray diffraction.
For these analyses, we have access to the following instruments:
Transmission electron microscopes:
FEI Tecnai F20 200kV FEG
Thermo Scientific GLACIOS 200kV FEG
Spectrometers:
950 MHz Bruker Avance III HD Liquid/Solid
850 MHz Bruker Avance III HD Liquid
600 MH Bruker Avance III HD Liquid/Solid
Crystallography and X-ray diffraction:
TTP-Labtech nanovolume crystallisation robot and visualisation robot
Access to the ESRF synchrotron
Methods – Protein/sample information
Circular Dichroism – % secondary structures
Fourier Transform Infrared Spectroscopy (FT-IR) – % secondary structures
Atomic Force Microscopy (AFM) – Protein orientation and transmembrane domain validation
Cryo-EM – Three-dimensional structure
X-ray diffraction – Three-dimensional structure
NMR
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Even
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