emerald bio nollert_pegs_draft_v4.3
DESCRIPTION
Presentation at PEGS2012 Boston, MATRANSCRIPT
May 3, 2012
8th Annual Protein Engineering Summit - PEGS 11/16/2012 Copyright 2012 Slide 1
Peter Nollert, Ph.D.
Chief Technology Officer
Emerald Bio
• Introduction Emerald Bio• Concepts & workflow• Multi-target parallel processing• Automated parallel protein purification
Parallel protein purification at the mg scale
Our Protein X-ray Crystallography Structure Output
11/16/2012 Slide 2
April 15: 482
PEGS 2012
SSGCID.org
Emerald is depositing 1 new structure every 1.7 days
SSGCID: the Most Productive
Structural Genomics Center in 2011
1.5 2.0 2.5 3.0Resolution (Å)
3/20/2012 PEGS 2012
MTPP: Multi-Target Parallel Processing
11/16/2012 Slide 4
Average Resolution: 2.3ÅAverage MolProbity: 94 percentile
MTPP and Quality
PEGS 2012
>100 structures delivered, >60 ligand bound13 different targets crystallized
Hundreds of Structures Require Thousands of Proteins
11/16/2012 Slide 5
5g
PEGS 2012
684 vector constructs220 different proteins purified9.7g = total protein produced
MPTT: Multi Target Parallel Processing
11/16/2012 6
131 E.coli132 Baculovirus
6 HEK293
231 Purifications
ConstructGeneration
X-ray StructureExpression Testing
ProteinSamples
Crystals
269 Constructs
14 targets as protein reagent16 targets selected 9 unique target structures
PEGS 2012
Emerald Bio’s : Protein Resource Operating System
7
Protein Crystals
Protein / ligand 3D co-structures
Fragment Based Drug Discovery
Ligand Optimization
Mechanism of Action
Intellectual Property
Protein Maker™
OptiMatrix Maker
Gene Composer
™
Process & Data Management
System
TOOLS
Pure Proteins
SAMPLES & INSIGHT
TARGET SPECIFIC PROCEDURE DEVELOPMENT
Expression Constructs
Recombinant Expression Systems
Formulations Purification
BIOLOGY & CHEMISTRY
Genes
Compounds
Target Info
E-screenBuilder
Applications: Antibody Purification
from up to 1 Liter load
• Biomarker discovery (immuno depletion)
• Parallel expression and purification testing
11/16/2012 8US Pat.# 6,818,060
High-throughput parallel protein purification with the Protein Maker
Parallel Protein Chromatography
• Carries out up to 24 purifications / run
• One step purification or two step purification (up to 12 samples)
• Uses standard 1-5 mL chromatography columns in parallel
• Unattended operation, up to 4 runs / day
• Small (1mL) to large (1L) load volumes
PEGS 2012
11/16/2012
Throughput:
Multiple targets & variants are
provided in batches
Approach: Parallel
chromatographic IMAC purification
of 24 target protein
9
Target variation:
Purification of internal deletion
variants; His-tagged
Analysis:
• SDS-PAGE
• Crystallization
Lysates Eluates Crystals
Protein Maker:Automated Parallel Protein Purification
PEGS 2012
Protein Maker: Parallel lysis scouting
Problem: Standard lysis buffer may not render all protein targets soluble.Solution: Test 12 different cell-lysis buffers followed by small scale (1 mL) IMAC purification
10
Sample: Cytochrome P450 insoluble in standard lysis bufferResult: of 12 lysis buffers 2 yield soluble CYP51A (CHAPS/bOG + 500 mM NaCl)
Lyse E.colicells in 12 different buffers
Ni-IMAC standard elution
protocol
SDS-PAGE analysis
Scale-up
cell lysis
Example
PEGS 2012Smith, E., et al. (2011). Acta Cryst.F , 67 (9), 1015-1021
11/16/2012
Protein Resources Operating System
11/16/2012 11
Protein Crystals
Protein / ligand 3D co-structures
Fragment Based Drug Discovery
Ligand Optimization
Mechanism of Action
Intellectual Property
Protein Maker
OptiMatrix Maker
Gene Composer
Process & Data Management
System
TOOLS
Pure Proteins
SAMPLES & INSIGHT
TARGET SPECIFIC PROCEDURE DEVELOPMENT
Expression Constructs
Recombinant Expression Systems
Formulations Purification
BIOLOGY & CHEMISTRY
Genes
Compounds
Target Info
E-screenBuilder
More Results:
• Emerald Bio @ PEGS trade show floor: booth# 62/63
• Email: [email protected]
11/16/2012 PEGS 2012 12