Bringing structural biology services through collaboration
1/ Collaboration key players Synthelis and ESRF SYNTHELIS / www. synthelis. com 2
Synthelis, a dynamic company Transforming academic expertise into relevant services Ø Spin-off company from University Joseph Fourier Grenoble - Incorporated in January 2011 Ø Bringing service offer to industries : development, production & characterization of difficult to express proteins (membrane & cytotoxic proteins) Proprietary technology and knowhow in cell-free protein expression SYNTHELIS / www. synthelis. com 3
Recognized expertise Already validated expertise with well-established customers Ø 33 client portfolio Ø 93% of protein successfully expressed Ø 77% with proof of activity Ø > 80 proteins successfully produced to date SYNTHELIS / www. synthelis. com 4
Structural biology cluster – Grenoble area SYNTHELIS / www. synthelis. com 5
ESRF in brief European Synchrotron Radiation Facility (ESRF) Ø Established in 1975 with the synchrotron project, capable of producing highly brilliant hard X-rays. Services Ø Beam time measurements Ø Experiment support by in-house experts Ø Industrial access • Simple and effective access to X-ray facilities and expertise • Specialized laboratory services • Technology transfer, instrument sales and licensing World reference for industrial and academic use of synchrotron radiation SYNTHELIS / www. synthelis. com 6
2/ Integrated offer From risk analysis to MAD/SAD structure solving SYNTHELIS / www. synthelis. com 7
Overall process* 10 to 14 months 1 week 5 - 6 months Se-Met expression Risk analysis Expression upscale 1 month Stability and activity assessment (TSA) 1– 2 month(s) Purification set-up & optimization 1 -2 month(s) Cristallogenesis Biophysics options If no crystals 1 week Diffraction test Data collection 2 weeks MAD/SAD structure solving If bad diffraction 1 -2 month(s) * Approximated timelines SYNTHELIS / www. synthelis. com 8
Risk analysis Service offer Ø Sequence analysis and structure prediction tools Ø Recommendation on which or how many construct(s) to be tested Ø Reporting and scientific meeting for GO/NO GO decision Sequence alignement Timeline 1 week Disorder prediction (Source: Thesis from E. Poudevigne) SYNTHELIS / www. synthelis. com 9
Se-Met expression & expression upscale Service offer Ø Two genetic construction design and production Ø Screening of cell-free expression conditions using Se-Met amino acids Ø Pre-purification by affinity Ø Western-Blot for identification and Coomassie Blue Staining gel for quantification Ø Expression scale up to produce mgs for characterization, purification and crystallogenesis. Timeline 5 - 6 months SYNTHELIS / www. synthelis. com 10
Stability and activity assessment (TSA) Service offer Ø For the 2 constructions in different conditions Ø Thermal shift assays to assess the activity and thermo-stability of the proteins Ø Reporting and scientific meeting for GO/NO GO decision Timeline 1 month TSA measurements SYNTHELIS / www. synthelis. com 11
Purification set-up & optimization Service offer Ø Purification with pre-determined protocol and concentration of sample up to 10 mg/ml if possible Ø Concentration and solubility tests Ø Reporting (Source: Thèse E. Poudevigne) SYNTHELIS / www. synthelis. com Timeline 1 -2 month(s) 12
Crystallogenesis Service offer Ø Crystallization set-up Ø Image Inspection/annotation Ø Screening of 1 protein samples up to 3 concentrations (one screening consisting of 5 -6 crystallization plates +1 refinement plate). Preparation of cryogenic solutions for up to best 5 hits per screen. Ø Reporting Timeline 1 -2 month(s) SYNTHELIS / www. synthelis. com 13
Biophysics options Service offer Ø Size and shape of macromolecules Ø Stoichiometry of macromolecular assemblies Ø Secondary and tertiary structure of macromolecules Ø Thermodynamic stability of macromolecules Ø Properties kinetic and thermodynamic interactions Timeline 2 -6 weeks depending on options SYNTHELIS / www. synthelis. com
Diffraction test Service offer Ø Test of crystal hits on synchrotron beamline Ø Feedback to crystallisation if further refinement required Timeline 1 week SYNTHELIS / www. synthelis. com 15
Crystal optimization Service offer Ø Manual production of crystals for data collection with optimized conditions from (5). Production of cryo-conditions, mounting and freezing crystals. Maximum duration of three months. Ø Manual crystal optimization Ø Consumables Ø Reporting Timeline 1 – 2 month(s) SYNTHELIS / www. synthelis. com 16
Structure solving and refinement Service offer Ø Protein structure determination – via molecular replacement or use of anomalous scattering Ø Subsequent refinement of structure Ø Reporting to level equivalent to PDB structure deposition Timeline 2 weeks for standard resolution SYNTHELIS / www. synthelis. com 17
3/ Why work together ? SYNTHELIS / www. synthelis. com 18
Benefits Ø Complementary expert teams Ø Proximity between the production site and the structure determination site Ø Single point-of-contact to access the whole value chain Ø Industry-driven service offer SYNTHELIS / www. synthelis. com 19
Your contact at SYNTHELIS : Alex Abadie 5 Avenue du Grand Sablon 38700 Grenoble - FRANCE Phone : +33 4 76 54 95 35 Mobile : +33 6 46 23 12 60 alex. abadie@synthelis. com www. synthelis. com SYNTHELIS / www. synthelis. com 20
Скачать презентацию Bringing structural biology services through collaboration 1
5b68bb76039c05a3a879fa6d7b2bf42b.ppt