Editing Proteome (section)

==Methods to study the proteome==
[[File:2D gel color coding.jpg|thumb|This image shows a two-dimensional gel with color-coded proteins. This is a way to visualize proteins based on their mass and isoelectric point.]]
{{Main|Proteomics}}

Analyzing proteins proves to be more difficult than analyzing nucleic acid sequences. While there are only 4 nucleotides that make up DNA, there are at least [[Proteinogenic amino acids|20 different amino acids that can make up a protein.]] Additionally, there is currently no known [[high throughput biology|high throughput]] technology to make copies of a single protein. Numerous methods are available to study proteins, sets of proteins, or the whole proteome. In fact, proteins are often studied indirectly, e.g. using computational methods and analyses of genomes. Only a few examples are given below.

===Separation techniques and electrophoresis===
[[Proteomics]], the study of the proteome, has largely been practiced through the separation of proteins by [[two dimensional gel electrophoresis]].  In the first dimension, the proteins are separated by [[isoelectric focusing]], which resolves proteins on the basis of charge.  In the second dimension, proteins are separated by [[Molecular mass|molecular weight]] using [[SDS-PAGE]].  The gel is [[Staining|stained]] with [[Coomassie brilliant blue]] or [[Silver staining|silver]] to visualize the proteins.  Spots on the gel are proteins that have migrated to specific locations.

===Mass spectrometry===
[[Image:ThermoScientificOrbitrapElite.JPG|thumb|An Orbitrap [[mass spectrometer]] commonly used in proteomics]]
{{main|Protein mass spectrometry|Mass spectrometry}}

[[Mass spectrometry]] is one of the key methods to study the proteome.<ref>{{cite journal|last=Altelaar|first=AF|author2=Munoz, J |author3=Heck, AJ |title=Next-generation proteomics: towards an integrative view of proteome dynamics.|journal=Nature Reviews Genetics|date=January 2013|volume=14|issue=1|pages=35–48|pmid=23207911|doi=10.1038/nrg3356|s2cid=10248311}}</ref> Some important mass spectrometry methods include Orbitrap Mass Spectrometry, [[MALDI]] (Matrix Assisted Laser Desorption/Ionization), and [[Electrospray ionization|ESI (Electrospray Ionization).]] [[Peptide mass fingerprinting]] identifies a protein by cleaving it into short peptides and then deduces the protein's identity by matching the observed peptide masses against a [[sequence database]].  [[Tandem mass spectrometry]], on the other hand, can get sequence information from individual peptides by isolating them, colliding them with a non-reactive gas, and then cataloguing the fragment [[Ion (physics)|ion]]s produced.<ref>{{cite journal |title=Mass-Spectrometry-Based Draft of the Human Proteome |url=https://www.jpt.com/literature/Mass-Spectrometry-Based-Draft-of-the-Human-Proteome |journal=[[Nature (journal)|Nature]] |volume=509 |issue=7502 |pages=582–7 |bibcode=2014Natur.509..582W |last1=Wilhelm |first1=Mathias |last2=Schlegl |first2=Judith |last3=Hahne |first3=Hannes |last4=Gholami |first4=Amin Moghaddas |last5=Lieberenz |first5=Marcus |last6=Savitski |first6=Mikhail M. |last7=Ziegler |first7=Emanuel |last8=Butzmann |first8=Lars |last9=Gessulat |first9=Siegfried |last10=Marx |first10=Harald |last11=Mathieson |first11=Toby |last12=Lemeer |first12=Simone |last13=Schnatbaum |first13=Karsten |last14=Reimer |first14=Ulf |last15=Wenschuh |first15=Holger |last16=Mollenhauer |first16=Martin |last17=Slotta-Huspenina |first17=Julia |last18=Boese |first18=Joos-Hendrik |last19=Bantscheff |first19=Marcus |last20=Gerstmair |first20=Anja |last21=Faerber |first21=Franz |last22=Kuster |first22=Bernhard |year=2014 |doi=10.1038/nature13319 |pmid=24870543 |s2cid=4467721 |access-date=2016-09-29 |archive-date=2018-08-20 |archive-url=https://web.archive.org/web/20180820005653/https://www.jpt.com/literature/Mass-Spectrometry-Based-Draft-of-the-Human-Proteome |url-status=dead }}</ref>

In May 2014, a draft map of the human proteome was published in ''[[Nature (journal)|Nature]]''.<ref>{{cite journal|last1=Kim|first1=Min-Sik|title=A draft map of the human proteome|journal=Nature|volume=509|doi=10.1038/nature13302|pmid=24870542|issue=7502|date=May 2014|pages=575–81|display-authors=etal|pmc=4403737|bibcode=2014Natur.509..575K}}</ref> This map was generated using high-resolution Fourier-transform mass spectrometry. This study profiled 30 histologically normal human samples resulting in the identification of proteins coded by 17,294 genes. This accounts for around 84% of the total annotated protein-coding genes.

=== Chromatography ===
Liquid [[chromatography]] is an important tool in the study of the proteome. It allows for very sensitive separation of different kinds of proteins based on their affinity for a matrix. Some newer methods for the separation and identification of proteins include the use of monolithic capillary columns, high temperature chromatography and capillary electrochromatography.<ref>{{Cite journal|last1=Shi|first1=Yang|last2=Xiang|first2=Rong|last3=Horváth|first3=Csaba|last4=Wilkins|first4=James A.|date=2004-10-22|title=The role of liquid chromatography in proteomics|journal=Journal of Chromatography A|series=Bioanalytical Chemistry: Perspectives and Recent Advances with Recognition of Barry L. Karger|volume=1053|issue=1|pages=27–36|doi=10.1016/j.chroma.2004.07.044|pmid=15543969|issn=0021-9673}}</ref>

=== Blotting ===
[[Western blot]]ting can be used in order to quantify the abundance of certain proteins. By using antibodies specific to the protein of interest, it is possible to probe for the presence of specific proteins from a mixture of proteins.

=== Protein complementation assays and interaction screens ===
[[Protein-fragment complementation assay]]s are often used to detect [[protein–protein interaction]]s. The [[Two-hybrid screening|yeast two-hybrid assay]] is the most popular of them but there are numerous variations, both used ''[[in vitro]]'' and ''[[in vivo]]''. Pull-down assays are a method to determine the protein binding partners of a given protein.<ref>{{Cite web|url=https://www.thermofisher.com/us/en/home/life-science/protein-biology/protein-biology-learning-center/protein-biology-resource-library/pierce-protein-methods/pull-down-assays.html|title=Pull-Down Assays - US|website=www.thermofisher.com|language=en|access-date=2019-12-05}}</ref>

=== Protein structure prediction ===
[[Protein structure prediction]] can be used to provide three-dimensional protein structure predictions of whole proteomes. In 2022, a large-scale collaboration between [[European Molecular Biology Laboratory|EMBL-EBI]] and [[DeepMind]] provided predicted structures for over 200 million proteins from across the tree of life.<ref>{{Cite journal |last=Callaway |first=Ewen |date=2022-07-28 |title='The entire protein universe': AI predicts shape of nearly every known protein |journal=Nature |language=en |volume=608 |issue=7921 |pages=15–16 |doi=10.1038/d41586-022-02083-2|pmid=35902752 |bibcode=2022Natur.608...15C |s2cid=251159714 |doi-access=free }}</ref> Smaller projects have also used protein structure prediction to help map the proteome of individual organisms, for example [https://www.isoform.io isoform.io] provides coverage of multiple protein isoforms for over 20,000 genes in the [[human genome]].<ref>{{Cite journal |last1=Sommer |first1=Markus J. |last2=Cha |first2=Sooyoung |last3=Varabyou |first3=Ales |last4=Rincon |first4=Natalia |last5=Park |first5=Sukhwan |last6=Minkin |first6=Ilia |last7=Pertea |first7=Mihaela |last8=Steinegger |first8=Martin |last9=Salzberg |first9=Steven L. |date=2022-12-15 |title=Structure-guided isoform identification for the human transcriptome |journal=eLife |volume=11 |pages=e82556 |language=en |doi=10.7554/eLife.82556|pmid=36519529 |pmc=9812405 |doi-access=free }}</ref>