This document provides an overview of common proteomics techniques. It describes proteomics as the study of proteins including their roles, structures, localization, interactions and other factors. The key techniques discussed include molecular techniques like DNA microarrays and yeast two-hybrid analysis, separation techniques like gel electrophoresis and chromatography, protein identification methods like mass spectroscopy and Edman sequencing, and protein structure determination methods like NMR, X-ray crystallography and computational prediction. The document provides examples and details of several of these techniques.
Introduction
Transcriptome analysis
Goal of functional genomics
Why we need functional genomics
Technique
1. At DNA level
2.At RNA level
3. At protein level
4. loss of function
5. functional genomic and bioinformatics
Application
Latest research and reviews
Websites of functional genomics
Conclusions
Reference
Functional proteomics, methods and toolsKAUSHAL SAHU
INTRODUCTION
HISTORY
DEFINITION
PROTEOMICS
FUNCTIONAL PROTEOMICS
PROTEOMICS SOFTWARE
PROTEOMICS ANALYSIS
TOOLS FOR PROTEOM ANALYSIS
DIFFERENTS METHODS FOR STUDY OF FUNCTIONAL PROTEOMICS
APLLICATIONS
LIMITATIONS
CONCLUSION
Yeast two-hybrid is based on the reconstitution of a functional transcription factor (TF) when two proteins or polypeptides of interest interact. Upon interaction between the bait and the prey, the DBD and AD are brought in close proximity and a functional TF is reconstituted upstream of the reporter gene.
Introduction
Transcriptome analysis
Goal of functional genomics
Why we need functional genomics
Technique
1. At DNA level
2.At RNA level
3. At protein level
4. loss of function
5. functional genomic and bioinformatics
Application
Latest research and reviews
Websites of functional genomics
Conclusions
Reference
Functional proteomics, methods and toolsKAUSHAL SAHU
INTRODUCTION
HISTORY
DEFINITION
PROTEOMICS
FUNCTIONAL PROTEOMICS
PROTEOMICS SOFTWARE
PROTEOMICS ANALYSIS
TOOLS FOR PROTEOM ANALYSIS
DIFFERENTS METHODS FOR STUDY OF FUNCTIONAL PROTEOMICS
APLLICATIONS
LIMITATIONS
CONCLUSION
Yeast two-hybrid is based on the reconstitution of a functional transcription factor (TF) when two proteins or polypeptides of interest interact. Upon interaction between the bait and the prey, the DBD and AD are brought in close proximity and a functional TF is reconstituted upstream of the reporter gene.
Secondary Structure Prediction of proteins Vijay Hemmadi
Secondary structure prediction has been around for almost a quarter of a century. The early methods suffered from a lack of data. Predictions were performed on single sequences rather than families of homologous sequences, and there were relatively few known 3D structures from which to derive parameters. Probably the most famous early methods are those of Chou & Fasman, Garnier, Osguthorbe & Robson (GOR) and Lim. Although the authors originally claimed quite high accuracies (70-80 %), under careful examination, the methods were shown to be only between 56 and 60% accurate (see Kabsch & Sander, 1984 given below). An early problem in secondary structure prediction had been the inclusion of structures used to derive parameters in the set of structures used to assess the accuracy of the method.
Some good references on the subject:
it will help you to understand how the protein microarrays are made, what are the different types and what all purposes they are used for. its very useful ppt
High throughput next generation sequencing and robust transcriptome analysis help with gene expression profiling, gene annotation or discovery of non-coding RNA.
Secondary Structure Prediction of proteins Vijay Hemmadi
Secondary structure prediction has been around for almost a quarter of a century. The early methods suffered from a lack of data. Predictions were performed on single sequences rather than families of homologous sequences, and there were relatively few known 3D structures from which to derive parameters. Probably the most famous early methods are those of Chou & Fasman, Garnier, Osguthorbe & Robson (GOR) and Lim. Although the authors originally claimed quite high accuracies (70-80 %), under careful examination, the methods were shown to be only between 56 and 60% accurate (see Kabsch & Sander, 1984 given below). An early problem in secondary structure prediction had been the inclusion of structures used to derive parameters in the set of structures used to assess the accuracy of the method.
Some good references on the subject:
it will help you to understand how the protein microarrays are made, what are the different types and what all purposes they are used for. its very useful ppt
High throughput next generation sequencing and robust transcriptome analysis help with gene expression profiling, gene annotation or discovery of non-coding RNA.
Proteomics, definatio , general concept, signficanceKAUSHAL SAHU
INTRODUCTION
GENERAL CONCEPT
WHY PROTEIOMIC NECESERY?
WHAT PROTEOMIC CAN ANSWER?
PRTEOMICS- ANALYSIS AND IDENTIFICATION OF PROTEIN
TWO-DIMENSIONAL SDS-PAGE
MASS SPECTROMETERS
SIGNIFICANCE OF STUDY AN ITS IMPORTANCE
APPLICATIONS
CHALLENGES
CONCLUSIONS
REFERENCES
Protein microarray Preparation of protein microarray Different methods of arr...naveed ul mushtaq
Protein microarray
Preparation of protein microarray
Different methods of arraying the proteins.FUNCTIONAL PROTEIN MICROARRAYSAnalytical microarrays:-
3.REVERSE PHASE PROTEIN MICROARRAY APPLICATIONS:-
Proteomics and its applications in phytopathologyAbhijeet Kashyap
Dear friends, I Abhijeet kashyap presenting the basics of proteomics to you all . Proteomics is the large-scale study of proteins, particularly their structures and functions.Proteomics helps in understanding the structure and function of different proteins as well as protein-protein interactions of an organism.
Proteomics: types, protein profiling steps etc.Cherry
Proteome is a set of proteins produced in an organism, system, or biological context or entire set of proteins that is, or can be, expressed by a genome, cell, tissue, or organism at a certain expressed time in a given set of condition. Proteomics is the study of all the proteins produced by a cell.
GENOMICS
Genomics is the study of all genes in an organism, also known as its genome. Genomics includes identifying the specific building blocks of all the genes in a cell, mapping their location in relation to the rest of the DNA, and studying the function of those genes or combination of those genes.
Types of Genomics :
1. Structural Genomics
2. Comparative Genomics
3.Functional Genomics
4. Epigenomics
5. Metagenomics
6. Pharmacogenomics
7. Mutation Genomics.
PROTEOMICS : (PROTEin in complement to genOME)
Proteomics is the study of proteome [Proteome is a protein molecule that interacts to give the cell its individual character]. Proteomics is a subset of functional genomics.
The proteome of a cell is all the proteins expressed by its genome. The proteome is of intense interest to investigators because proteins are the major functional components of the cell.
Proteomics is the study of proteins in order to revolutionize the understanding of cell behaviour and disease.
1. It studies the translation of process of RNA into proteins as well as the overall process of DNA into proteins.
2. It studies the diseases through proteins because disease process manifest themselves at the level of protein activity.
3. Most drugs act by targeting proteins or protein receptors, so Proteomics is important in new generation of drugs.
4. Proteins are more complex than genes because they can be modified after formation.
5. Proteomics is the qualitative and quantitative comparison of proteomes under different conditions to further unravel biological processes.
6. Proteomics can use analysis techniques to determine all of the post translational modifications that proteins undergo and therefore determine what makes a diseased or mutant protein different from a normal protein.
Proteins are fundamental components of all living cells. Proteins help us digest our food, fight infections, control body chemistry, keep our bodies function smoothly. Identifying a proteins’ shape or structure is key to understanding its biological function and its role in health and disease.
There are different strategies bacterial cells use to survive. Differentiation can be occasionally one of them. Although differentiation can occur in the bacterial life cycle, it is a strategy to adapt themselves to harsh environments.
Deep Behavioral Phenotyping in Systems Neuroscience for Functional Atlasing a...Ana Luísa Pinho
Functional Magnetic Resonance Imaging (fMRI) provides means to characterize brain activations in response to behavior. However, cognitive neuroscience has been limited to group-level effects referring to the performance of specific tasks. To obtain the functional profile of elementary cognitive mechanisms, the combination of brain responses to many tasks is required. Yet, to date, both structural atlases and parcellation-based activations do not fully account for cognitive function and still present several limitations. Further, they do not adapt overall to individual characteristics. In this talk, I will give an account of deep-behavioral phenotyping strategies, namely data-driven methods in large task-fMRI datasets, to optimize functional brain-data collection and improve inference of effects-of-interest related to mental processes. Key to this approach is the employment of fast multi-functional paradigms rich on features that can be well parametrized and, consequently, facilitate the creation of psycho-physiological constructs to be modelled with imaging data. Particular emphasis will be given to music stimuli when studying high-order cognitive mechanisms, due to their ecological nature and quality to enable complex behavior compounded by discrete entities. I will also discuss how deep-behavioral phenotyping and individualized models applied to neuroimaging data can better account for the subject-specific organization of domain-general cognitive systems in the human brain. Finally, the accumulation of functional brain signatures brings the possibility to clarify relationships among tasks and create a univocal link between brain systems and mental functions through: (1) the development of ontologies proposing an organization of cognitive processes; and (2) brain-network taxonomies describing functional specialization. To this end, tools to improve commensurability in cognitive science are necessary, such as public repositories, ontology-based platforms and automated meta-analysis tools. I will thus discuss some brain-atlasing resources currently under development, and their applicability in cognitive as well as clinical neuroscience.
Multi-source connectivity as the driver of solar wind variability in the heli...Sérgio Sacani
The ambient solar wind that flls the heliosphere originates from multiple
sources in the solar corona and is highly structured. It is often described
as high-speed, relatively homogeneous, plasma streams from coronal
holes and slow-speed, highly variable, streams whose source regions are
under debate. A key goal of ESA/NASA’s Solar Orbiter mission is to identify
solar wind sources and understand what drives the complexity seen in the
heliosphere. By combining magnetic feld modelling and spectroscopic
techniques with high-resolution observations and measurements, we show
that the solar wind variability detected in situ by Solar Orbiter in March
2022 is driven by spatio-temporal changes in the magnetic connectivity to
multiple sources in the solar atmosphere. The magnetic feld footpoints
connected to the spacecraft moved from the boundaries of a coronal hole
to one active region (12961) and then across to another region (12957). This
is refected in the in situ measurements, which show the transition from fast
to highly Alfvénic then to slow solar wind that is disrupted by the arrival of
a coronal mass ejection. Our results describe solar wind variability at 0.5 au
but are applicable to near-Earth observatories.
Nutraceutical market, scope and growth: Herbal drug technologyLokesh Patil
As consumer awareness of health and wellness rises, the nutraceutical market—which includes goods like functional meals, drinks, and dietary supplements that provide health advantages beyond basic nutrition—is growing significantly. As healthcare expenses rise, the population ages, and people want natural and preventative health solutions more and more, this industry is increasing quickly. Further driving market expansion are product formulation innovations and the use of cutting-edge technology for customized nutrition. With its worldwide reach, the nutraceutical industry is expected to keep growing and provide significant chances for research and investment in a number of categories, including vitamins, minerals, probiotics, and herbal supplements.
1. A review to proteomics common methods
TECHNIQUES IN PROTEOMICS
2. Definitions
o Proteome: The set of all expressed proteins in a cell, tissue,
or organism.
o Proteomics: A science that focuses on the study of proteins
(their roles, their structures, their localization, their
interactions, and other factors).
o SO there should be methods to understand
these items!
3. Proteomics techniques
o Molecular techniques
o Separation techniques
o Protein Identification techniques
o Protein Structure techniques
4. Molecular techniques
I. DNA Microarrays or Gene Chips
II. Differential Display*
III. Northern/Southern Blotting
IV. RNAi (small RNA interference)
V. Serial Analysis of Gene Expression (SAGE)*
VI. Yeast two-hybrid analysis
5. SAGE
o a technique used to produce a snapshot of the messenger
RNA population in a sample of interest in the form of small
tags that correspond to fragments of those transcripts,
developed by Dr. Victor Velculescu at the Oncology Center
of Johns Hopkins University and published in 1995.
o Although SAGE was originally conceived for use in cancer
studies, it has been successfully used to describe
the transcriptome of other diseases and in a wide variety of
organisms.
6.
7.
8. Differential Display (DD-PCR)
o Since its invention in the early 1990s, differential display has
become one of the most commonly used techniques for
identifying differentially expressed genes at the mRNA level.
o The essence of the method is to amplify messenger RNA 3'
termini using a pair of anchored oligo-dT primer and a short
primer with an arbitrary sequence.
o The amplified cDNAs labeled with radioisotope are then
distributed on a denaturing polyacrylamide gel and visualized
by autoradiography.
o In the mid-2000s, differential display and RNAse protection
assay were superseded by DNA Microarrays.
9. Proteomics techniques
o Molecular techniques
o Separation techniques
o Protein Identification techniques
o Protein Structure techniques
10. Separation techniques
I. 1D Slab Gel Electrophoresis
II. 2D Gel Electrophoresis (SDS-PAGE/IEF)
III. Capillary Electrophoresis
IV. Chromatography (HPLC ,SEC, IEC, RP, Affinity, etc.)
V. Protein Chips (Protein microarray)*
11. Protein array(protein chip)
o They are modeled after DNA microarrays, in 2000 at Harvard University
o The success of DNA microarrays in large-scale genomic experiments
inspired researchers to develop similar technology to enable large-scale,
high-throughput proteomic experiments.
o Protein chips enable researchers to quickly and easily survey the entire
proteome of a cell within an organism.
o Applications include:
- identifying biomarkers for diseases,
- investigating protein-protein interactions,
- testing for the presence of a protein(i.e. Ab) in a sample.
13. 1.Analytical microarrays (capture arrays)
o Used to understand:
- expression levels,
- binding affinities and specificities,
- response of the cells to a particular factor,
- identification and profiling of diseased tissues.
14. 2. Functional protein microarrays (target
protein arrays)
o Immobilised purified proteins are used to:
- identify protein-protein/DNA/RNA/PL/SM,
- assay enzymatic activity.
o They differ from analytical arrays in that they contain full-
length functional proteins.
15. 3. Reverse phase protein microarray
(RPPA)
o involve complex samples, such as tissue lysates probed with
antibodies against the target protein of interest.
o These antibodies are typically detected
with chemiluminescent, fluorescent or colorimetric assays.
o Used to: determination of the presence of altered proteins or
other agents as a result of disease.
Specifically, post-translational modifications, which are
typically altered as a result of disease.
16. Proteomics techniques
o Molecular techniques
o Separation techniques
o Protein Identification techniques
o Protein Structure techniques
17. Protein Identification techniques
I. Edman sequencing
II. Microsequencing
III. Mass spectroscopy
o Sequencing done for:
1. Protein's amino acid three-dimensional structure.
2. Sequence comparisons among analogous proteins
protein function and reveal evolutionary relationships.
3. Many inherited diseases are caused by mutations leading to an
amino acid change in a protein.
18. Edman sequencing
o Mechanism:
-The peptide to be sequenced (50 aa) is adsorbed onto a solid surface -
one common is glass fibre coated with polybrene, a cationic polymer.
-The Edman reagent, phenylisothiocyanate (PITC) , is added to the
adsorbed peptide, together with a mildly basic buffer solution. This reacts
with the amine group of the N-terminal amino acid.
-The terminal amino acid can then be selectively detached by the addition
of anhydrous acid. It can be washed off and identified by chromatography,
and the cycle can be repeated.
o Limitations:
-it will not work if the N-terminal amino acid has been chemically
modified.
-separate procedure to determine the positions of disulfide bridges
needed.
21. Proteomics techniques
o Molecular techniques
o Separation techniques
o Protein Identification techniques
o Protein Structure techniques
22. Protein Structure techniques
I. NMR(Nuclear magnetic resonance spectroscopy)
II. X-ray crystallography
III. Computational prediction
NMR X-ray crystallography
23. NMR and X-ray crystallography help
o the two methods are very basic and critical in the field of protein
structure determination.
o The strengths and weaknesses of one of the two methods
fortunately supplement the holes and gaps in the other method to
make it possible that different kind of important data for a
structural question can be answered with the parallel or
supplemental application of the two methods.
o There may be several instances when only one method could be
used.
o If the two methodologies had the same basics, principles,
strengths and weaknesses we may be not capable to solve lots of
structural problems.
o We should thank this possibility to the technical development
which could catch different biophysical principles.
25. Computational prediction
o in theory, a protein structure can solved computationally as it folds into a
3D structure to minimizes its potential energy.
o ab initio/de novo folding methods: not practical (yet) due to its high
computational complexity. These procedures tend to require vast
computational resources, and have thus only been carried out for tiny
proteins.
o Comparative modeling methods: uses previously solved structures as
starting points, or templates.
a. Protein threading – make structure prediction through identification
of “good” sequence-structure fit
b. Homology modeling – identification of homologous proteins through
sequence alignment; structure prediction through placing residues
into “corresponding” positions of homologous structure models
28. The goal
“The ultimate goal of systems biology is the integration of data
from these observations into models that might, eventually,
represent and simulate the physiology of the cell.”