Human Proteome Project?
Màster en bioquímica, biologia molecular i biomedicina
Mòdul 4: Genòmica i Proteòmica
Núria Colomé Calls
After the Human Genome Project, the Human
Proteome Project seems a natural succesor
John Bergeron president of the Human Proteome Organisation (HUPO) and a
group of leading researchers propose a large-scale assault on the human
proteome.
This group plans to consult with
the proteomics community for
the first time at HUPO’s
Amsterdam World Congress in
August.
Human Proteome Project is now feasible
because:
-The number of protein coding genes is around
21000 making the scale of human proteomics
more manageable
Human Proteome Project would reveal:
Which proteins are present in each tissue
Where is located each protein in the cell
Protein-protein interactions
Invaluable information with clinical utility
Difficulties:
There are around 200 types of human cells that manufacture proteins at differing
levels and different moments in time.
Protein-coding genes in the body can make tens of different versions of a protein.
Each of these can be modified by addition of chemical groups.
The researchers’ plan is to use different experimental approaches:
Mass spectrometry : To identify proteins and their quantities
Generate antibodies to each protein: to show its location in tissues and cells
Systematic Identification protein-protein interactions
Bioinformatics tools design: to pool and make accessible the data
Proteins Identification
Initial assay: HUPO Human Plasma Proteome Project
(HPPP)
Proteins Identification
Initial assay: HUPO Human Plasma Proteome Project (HPPP)
2002 the Plasma Proteome Project was initiated.
The aim is the systematic characterization of circulating proteins in health and
disease to facilitate development of biomarkers for prevention, diagnosis and
therapy of different diseases
There were 35 collaborating laboratories and multiple analytical groups that were
provided with 1.0mL of serum and plasma by three different methods of
anticoagulation for plasma.
Different strategies to analyse the samples were used:
- Depletion or not of abundant proteins: albumin and/or
IgG and Top 6
- Fractionation of intact proteins: 2DE, LC or IEF methods
- Peptides separation: Cation exchange and/or Reverse
phase
- Multiple mass spectrometry instruments: ESI MS/MS,
MALDI MS and FT-ICR-MS
- Varied search algorithms and databases: PepMiner,
SEQUEST, MASCOT and VIPER
Abundance and dynamic range of plasma proteins
Proteins Identification
Initial assay: HUPO Human Plasma Proteome Project (HPPP)
Results
18 laboratories submitted a total of 42306 protein identifications.
This initial assay has provided a core data set of 3020 proteins based on two or more peptide, without redundant
entries and one representative protein among multiple proteins (homologs and isoforms)
The laboratories that reported more results had used a depletion method, peptide chromatographic separation
and MSMS instruments
Proteins Identification
Initial assay: HUPO Human Plasma Proteome Project (HPPP)
Comments
There is a high variability on the results depending on sample preparation, fractionation
and detection methods. Probably complementary.
These results lay a foundation for development and validation of circulating protein
biomarkers in health and disease and achieve quantitative comparisons across
specimens, not just compositional analyses
There is a Human Proteome contribution to human genome: 185 proteins are not
associated with genes EnsEMBL and 118 peptides were identified as highly probable
matches to ORFs in the human genome not previously known to have protein products
Generate antibodies to each protein to show its location
Protein Atlas: Part of the HUPO Human Antibody Initiative (HAI)
Generate antibodies to each protein to show its location
Protein Atlas: Part of the HUPO Human Antibody Initiative (HAI)
The human protein atlas shows expression and localization of proteins in a large variety of normal
human tissues, cancer tissues and cell lines with the aid of immunohistochemistry (IHC) images.
Subcellular
localization
results for 769
antibodies in
three different
cells lines
Atlas content:
3014 antibodies and
2,940,744 images.
Generate antibodies to each protein to show its location
Protein Atlas: Part of the HUPO Human Antibody Initiative (HAI)
HER2 Search with links to Uniprot, NCBI and EnsEMBL databases and antibody results
Generate antibodies to each protein to show its location
Protein Atlas: Part of the HUPO Human Antibody Initiative (HAI)
ERBB2 expression profiles for
HPA001383 antibody in Normal
tissues, Cancer tissues and Cell
lines
Generate antibodies to each protein to show its location
Protein Atlas: Part of the HUPO Human Antibody Initiative (HAI)
ERBB2 expression profiles images for HPA001383 antibody in Breast Cancer Tissue
Generate antibodies to each protein to show its location
Protein Atlas: Part of the HUPO Human Antibody Initiative (HAI)
Cell Location of BCAN protein:
Immunofluorescent (IF) images generated with confocal microscopy
Identification of Protein-protein interactions
Large-scale mapping of human protein–protein interactions by mass
spectrometry
Identification of Protein-protein interactions
Large-scale mapping of human protein–protein interactions by mass spectrometry
This study presents the first large-scale study of protein–protein interactions in human cells using a
Immunoprecipitation high-throughput mass spectrometry (IP-HTMS).
The study maps protein interactions for 338 bait proteins that were selected based on known or suspected
disease and functional associations. 6463 interactions were found
The data set provides for both recapitulation of known complexes and discovery of new interactions and
complexes.
Bait to bait conectivity map of bait proteins
Focused view of selected bait-bait networks
Bioinformatics
Bioinformatics
Develop a unique reservoire to pool and make accessible all the data
generated since now
Exist different datasets for human
plasma or serum
Healthy Human Individual’s Integrated Plasma proteome
(HIP2)
This database has been developed to be the comprehensive collection
of healthy human plasma proteins, it contains mappings of supporting
peptide evidence from several high quality and high-throughput mass
spectrometry (MS) experimental datasets. Users can search for plasma
protein/peptides annotations, peptide/protein alignments, and
experimental/sample conditions.
Bioinformatics
Other repositories:
PRIDE: PRoteomics IDEntifications database is a centralized, standards compliant, public data
repository for proteomics data. PRIDE is able to capture details of post-translational modifications.
This database contain: 3,745 Experiments,726,086 Identified Proteins , 4,618,998 Identified Peptides,
466,837 Unique Peptides and 6,446,100 Spectra
Peptide Atlas: PeptideAtlas is a multi-organism, publicly accessible compendium of peptides
identified in a large set of tandem mass spectrometry proteomics experiments
Protein Atlas: The human protein atlas shows expression and localization of proteins
Human Protein Atlas
Opinions
“The whole effort could take a decade and would involve more labs than the human
genome project”. Bergeron, Nature News
“Rather than analyse the proteome of one chromosome, it may be better to tackle the
proteome of mitochondria or the cell membrane because it would reveal more about
biology and diseases”. Steven Carr, Nature News
“Proteome Project would involve many labs, with inevitable issues about data-sharing
and competition”. Michael Washburn, Nature News
“The project may have a better chance of success with stronger focus on diseases
such as cancer, you need to show clinical utility of the approach before launching a
mega-project”. Sudhir Srivastava, Nature News
References
Overview of the HUPO Plasma Proteome Project: Results from the pilot phase with 35 collaborating laboratories
and multiple analytical groups, generating a core dataset of 3020 proteins and a publicly-available database.
Gilbert S. Omenn et al., Proteomics 2005, 5, 3226–3245
Data management and preliminary data analysis in the pilot phase of the HUPO Plasma Proteome Project. Marcin
Adamski et al., Proteomics 2005, 5, 3246–3261
Large-scale mapping of human protein–protein interactions by mass spectrometry. Rob M Ewing, Molecular
Systems Biology 2007,3:89, 1-17
HIP2: An Online Database of Human Plasma Proteins from Healthy Individuals. Sudipto Saha, BMC Medical
Genomics 2008, 1:12
http://www.nature.com/news/index.html
http://www.proteinatlas.org/
http://www.hupo.org/
http://bio.informatics.iupui.edu/HIP2/
http://www.ebi.ac.uk/pride/init.do