role of proteomics in target discovery and validation
1 target of drug action
2 proteomics
3 facts about proteins
4 post translational modification
5 additional modification
6 methods of studying proteins
7 hybrid technologies
3. Earliest drugs were
discovered with no
knowledge of their
mechanism or site
of action.
Modern drugs
inventions started
with a statement
of target.
Target is a certain protein or a pathway play a critical role in
pathogenesis of a disease which is altered by a drug to produce
its effects.
4. Proteomics is large scale study of proteins. The
term proteomics was coined in 1997,in
analogy to genomics, the study of genome.
It is more complicated then genomics because
an organismโs genome is constant, whereas
proteomes differ from cell to cell and from
time to time.
Proteomics confirms presence of the protein
and measure its quantity.
5. Proteins are polymer chains made of amino acids linked together by
peptide bonds. During human digestion, proteins are broken down
to smaller polypeptide chains via hydrochloric acid and protease
actions.
6. Following functions of proteins in human body:
โข Structural component of cells
โข Precursor to nucleic acid
โข Co-enzymes
โข Hormones
โข Immune response
โข Cellular repair mehanism.
7. Proteins undergoes a vide variety of chemical
modification after translation.
Phosphorylation :
Addition of phosphate to many enzymes and
structural proteins in process of cell signaling
e.g. phosphorylation of serine and threonine
mediated by serine-threonine kinase. and of
tyrosine mediated by tyrosine kinase โ cause
a protein to become a targetโฆ
8. In addition to phosphorylation some other
modifications are:
๏ถ Methylation
๏ถ Acetylation
๏ถ Oxidation
๏ถ Glycosylation
๏ถ Nitrosylation
Some proteins
undergoes all these
modification
depending upon
time.
9. A cell may make different sets of proteins at
different times or different conditions.
e.g.
Cell
development
Cellular
differentiation
Cell cycle
10. Generally proteins may be detected by using
either antibody(immunoassay) or mass
spectrometry, for complex biological samples
a biochemical separation need to be used
before detection step.
๏ฑ Immunoassay
๏ฑ Anti-body free protein detection
๏ฑ Hybrid technologies
๏ฑ Current research methodologies
๏ฑ High-throughput protein technologies
11. These are most common tool used by
molecular biologist today.
๏ฝ ELISA-enzyme linked immunosorbent assay
can be used for detection and quantification
of proteins.
๏ฝ WESTERN-BLOT used for detection and
quantification of individual proteinโฆ
๏ฝ Modified proteins may be studied by
developing an antibody specific to that
modification โฆ
12. ๏ฝ Disease detection at molecular level is a
revolution of early diagnosis & treatment โฆ
13. Methods have been developed for protein
identification without use of antibody. they
offers some advantages such as:
๏ฝ They determine the sequence of protein or
peptide
๏ฝ Also identify proteins for which no antibody
is present.
14. More recent implemented method used mass
spectrometry-based technique.
๏Matrix-assisted laser desorption/
ionization.(MALDI)
๏Electro spray ionization (ESI)
Robotic preparation of MALDI mass
Spectrometry sample
15. There are several hybrid technologies that use
antibody and then perform mass
spectrometric analysis for identification and
quantification
e.g. MSIA(mass spectrometric immunoassay)
Developed by Randall Nelson in 1995
16. Fluorescence two-dimensional differential gel
electrophoresis.
Comparative proteomic analysis may reveal the
role of proteins in complex biological system,
including reproduction.
e.g. treatment with insecticide triazophos
cause increase in content of planthopper โฆ
17. Large scale study of proteins including:
โข Mass spectrometry-based methods &
โข Micro arrays
Mass spectrometry and protein profiling
Two mass spectroscopy based methods are
currently used:
๏ฝ Two dimensional electrophoresis to separate
proteins from different samples and then
identified by mass spectrometry.
18. ๏ฝ The second approach uses stable isotope
tags to differentially label proteins โฆ
Reverse-phased protein microarrays
The technology merges laser capture
microdissection (LCM) with microarray
technology to produce reverse-phased
microarrays.
19. ๏ฝ Transcription of gene gives only rough
estimate of translation into a protein. mRNA
produced in abundance may be degraded
rapidly resulting in small amount of protein.
๏ฝ Post-translation modifications affects protein
activities e.g. some proteins are not active
until they become phosphorylated.
๏ฝ Many transcript give rise to more than one
protein through post-translational
modification.
20. ๏ฝ Many proteins form complexes with other
proteins or RNA molecules, and only function
in the presence of these other proteins.
๏ฝ REPRODUCIBILITY
It is the simultaneous elution of many more peptides
than mass spectrometers can measure. This causes
considerable variations between laboratories.
21. 1) Goodman & Gilmanโs โThe Pharmacological
Basis of Therapeutics โ12th edition, page
no.6
2) WIKIPEDIA โ Proteomics.
