Transcript Folie 1 - JPT Peptide Technologies

Advanced Library Generation
Tobias Knaute / Johannes Zerweck
Peptide Arrays
JPT Peptide Technologies, Berlin
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What´s the Problem?
• The peptide library generation for determining
antibody / antigen interactions gets complex if
many proteins are involved
• Post-translational modifications increase
complexity even more
• Number of peptides is limited due to
experimental setup (max. 7000 peptides per
experiment)
• Problem: How to generate the most efficient
library concerning coverage and content
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Conventional Method
Fixed Overlap
Overlap = Length - Offset
Generation of overlapping peptides
based on primary structure
C
N
MGARASVLSGGELDR
RASVLSGGELDRWEK
VLSGGELDRWEKIRL
GGELDRWEKIRLRPG
LDRWEKIRLRPGGKK
...
PRO:
CON:
• Easy to understand & implement
• Sufficient for simple screenings
and epitope mappings
• Insufficient for many homologous
sequences
• Could lead to bloated peptide lists
• Coverage not adjustable
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Advanced Method
Dynamical Overlap
• Algorithm enabling optimal coverage of protein
sequences by peptide libraries
• Task 1: find optimum between number of peptides and
coverage
• Task 2: generate peptide-score according to frequency
of motifs (e.g. in other homologous sequences)
• Task 3: ability to inject fixed sequences (e.g. from
previous projects)
• Task 4: ability to adjust coverage and/or number of
peptides to specific needs
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Example
Protein 1
Protein 2
Task:
- 2 homologous sequences
- make peptide scan
- peptide length 15
- maximum overlap 11
- make sure every residue is covered by at least 3 peptides
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Example - Convential Scan
Convential scan
Fixed overlap 15/11
Peptides of protein 1
Peptides
Protein 1
Protein 2
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Example - Convential Scan
Convential scan
Fixed overlap 15/11
Peptides of protein 2
Peptides
Protein 1
Protein 2
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Example - Convential Scan
Convential scan
Fixed overlap 15/11
All individual peptides,
mapped to protein 1
 Several peptides
from protein 2 fit into
sequence of protein 1
Peptides
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5
6
7 8 7
7
7
5
4
Protein 1
Protein 2
Redundancy introduced by homology.
Number of peptides
covering specific amino acid
in protein sequence
 amino acid coverage
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Example - Dynamical Overlap
All peptides, mapped to
protein 1
 Redundant peptides are
rejected by algorithm
overlap 8
Peptides
3
3
4 4 4
4
4
4
3
Protein 1
Protein 2
Overlap adjustable by coverage constraint.
Amino acid coverage
is optimized
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Case Study: HIV1 ENV gp160
24 Sequences of Group M Subtype B
Task:
Minimum coverage:
3 peptides/residue
Length: 15
Max. overlap: 11
highly conserved
region
highly variable
region
3283 peptides needed for traditional method
2303 peptides needed for advanced method
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Case Study: HIV1 ENV gp160
Residue Coverage Map
Conventional
3283 peptides
mean density:
6.7 peptides/residue
Advanced
2303 peptides
mean density:
3.7 peptides/residue
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Summary
• Algorithm to build optimal libraries developed
• Coverage / number of peptides can be adjusted
• Elimination of redundant motifs
• Addresses diversity, best suited for proteome-wide
sequence alignments in complex libraries
• Applicable to PepStarTM microarrays, PepTrackTM
Libraries, PepMixTM peptide pools ...
• Based on the input of protein sequences, JPT will develop
an optimized library for seroprofiling or immune monitoring
experiments
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
JPT Peptide Technologies
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History, Quality Assurance & Business Model
Quality Assurance
History
2002 Quality Management System
1994 Jerini Spin-Off from Charité Clinics Berlin
2004 DIN ISO 9001:2000 Certification
2002 Jerini Peptide Technologies - Business Unit
Since 2008 GCLP Compliance Audits (NIH, IAVI, BCM,
2004 JPT Peptide Technologies GmbH
Cellmedica, Genticel...)
2005 JPT Inc. in Boston, MA, USA
2010 DIN ISO 9001:2008 Certification
2006 New JPT Facilities in Berlin
2009 Acquisition by BioNTech AG
2011 50 Employees, 13 PhD‘s
Offices & Reps
Business Model
Berlin, Boston, Denver, Brussels
1500 sqm wet lab and 1200 sqm office space
Capabilities for organic & biological laboratory work
Approved S2 lab for infectious patient samples
Cleanroom microarray capabilities
Business develoment & marketing
http://shop.jpt.com
http://jpt.com
http://rnd.jpt.com
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Major Application Fields of Peptides
Immunology
• Vaccine development
• B- and T-cell epitope discovery
• Diagnostic tools for infectious, autoimmune
diseases, allergies & cancer
• Immune monitoring & immune therapies
Proteomics & Enzyme Profiling
• Enzyme profiling and protein/protein interaction studies
• Quantification of protein expression levels
• Biomarker discovery
Drug Discovery
• Biomedical screening
• Peptide therapeutics
• SAR, structure activity relationship studies
Feel free to discuss your specific needs with our experts !
JPT Peptide Technologies GmbH
Volmerstrasse 5
12489 Berlin, Germany
T +49-30-6392-5500
X +49-30-6392-5501
[email protected]
www.jpt.com
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for Custom Products & Services
for Peptide Related Collaboration
Projects
for Catalog Products
http://jpt.com