Editing Protein isoform

{{Short description|Forms of a protein produced from different genes}}
[[File:DNA alternative splicing.gif|thumb|Protein A, B and C are isoforms encoded from the same gene through [[alternative splicing]].|420x420px]]
A '''protein isoform''', or "'''protein variant'''",<ref name=":4" /> is a member of a set of highly similar [[proteins]] that originate from a single [[gene]] and are the result of genetic differences.<ref>{{cite journal | vauthors = Schlüter H, Apweiler R, Holzhütter HG, Jungblut PR | title = Finding one's way in proteomics: a protein species nomenclature | journal = Chemistry Central Journal | volume = 3 | pages = 11 | date = September 2009 | pmid = 19740416 | pmc = 2758878 | doi = 10.1186/1752-153X-3-11 | doi-access = free }}</ref> While many perform the same or similar biological roles, some isoforms have unique functions. A set of protein isoforms may be formed from [[Alternative splicing|alternative splicings]], variable [[Promoter (genetics)|promoter]] usage, or other [[Post-transcriptional modification|post-transcriptional modifications]] of a single gene; [[post-translational modification]]s are generally not considered. (For that, see [[Proteoform]]s.) Through [[RNA splicing]] mechanisms, [[mRNA]] has the ability to select different protein-coding segments ([[Exon|exons]]) of a gene, or even different parts of exons from RNA to form different mRNA sequences. Each unique sequence produces a specific form of a protein.

The discovery of isoforms could explain the discrepancy between the small number of protein coding regions of genes revealed by the [[human genome project]] and the large diversity of proteins seen in an organism: different proteins encoded by the same gene could increase the diversity of the [[proteome]]. Isoforms at the RNA level are readily characterized by [[Complementary DNA|cDNA]] transcript studies. Many human genes possess confirmed [[alternative splicing]] isoforms. It has been estimated that ~100,000 expressed sequence tags ([[Expressed sequence tag|ESTs]]) can be identified in humans.<ref name=":4">{{cite journal | vauthors = Brett D, Pospisil H, Valcárcel J, Reich J, Bork P | title = Alternative splicing and genome complexity | journal = Nature Genetics | volume = 30 | issue = 1 | pages = 29–30 | date = January 2002 | pmid = 11743582 | doi = 10.1038/ng803 | s2cid = 2724843 }}</ref> Isoforms at the protein level can manifest in the deletion of whole domains or shorter loops, usually located on the surface of the protein.<ref>{{Cite book | doi = 10.1002/9783527636778.ch54| chapter = Structure Prediction for Alternatively Spliced Proteins| title = Alternative pre-mRNA Splicing| pages = 582| year = 2012| last1 = Kozlowski | first1 = L. | last2 = Orlowski | first2 = J. | last3 = Bujnicki | first3 = J. M. | isbn = 9783527636778}}</ref>

== Definition ==
One single gene has the ability to produce multiple proteins that differ both in structure and composition;<ref name=":0">{{cite journal | vauthors = Andreadis A, Gallego ME, Nadal-Ginard B | title = Generation of protein isoform diversity by alternative splicing: mechanistic and biological implications | journal = Annual Review of Cell Biology | volume = 3 | issue = 1 | pages = 207–42 | date = 1987-01-01 | pmid = 2891362 | doi = 10.1146/annurev.cb.03.110187.001231 }}</ref><ref name=":1">{{cite journal | vauthors = Breitbart RE, Andreadis A, Nadal-Ginard B | title = Alternative splicing: a ubiquitous mechanism for the generation of multiple protein isoforms from single genes | journal = Annual Review of Biochemistry | volume = 56 | issue = 1 | pages = 467–95 | date = 1987-01-01 | pmid = 3304142 | doi = 10.1146/annurev.bi.56.070187.002343 }}</ref> this process is regulated by the [[alternative splicing]] of mRNA, though it is not clear to what extent such a process affects the diversity of the human proteome, as the abundance of mRNA transcript isoforms does not necessarily correlate with the abundance of protein isoforms.<ref>{{cite journal | vauthors = Liu Y, Beyer A, Aebersold R | title = On the Dependency of Cellular Protein Levels on mRNA Abundance | journal = Cell | volume = 165 | issue = 3 | pages = 535–50 | date = April 2016 | pmid = 27104977 | doi = 10.1016/j.cell.2016.03.014 | doi-access = free | hdl = 20.500.11850/116226 | hdl-access = free }}</ref> Three-dimensional protein structure comparisons can be used to help determine which, if any, isoforms represent functional protein products, and the structure of most isoforms in the human proteome has been predicted by [[AlphaFold]] and publicly released at [https://www.isoform.io isoform.io]. <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> The specificity of translated isoforms is derived by the protein's structure/function, as well as the cell type and developmental stage during which they are produced.<ref name=":0" /><ref name=":1" /> Determining specificity becomes more complicated when a protein has multiple subunits and each subunit has multiple isoforms.

For example, the '''[[AMP-activated protein kinase|5' AMP-activated protein kinase]]''' (AMPK), an enzyme, which performs different roles in human cells, has 3 subunits:<ref name=":2">{{cite journal | vauthors = Dasgupta B, Chhipa RR | title = Evolving Lessons on the Complex Role of AMPK in Normal Physiology and Cancer | language = English | journal = Trends in Pharmacological Sciences | volume = 37 | issue = 3 | pages = 192–206 | date = March 2016 | pmid = 26711141 | pmc = 4764394 | doi = 10.1016/j.tips.2015.11.007 }}</ref>
* α, catalytic domain, has two isoforms: α1 and α2 which are encoded from [[PRKAA1]] and [[PRKAA2]]
* β, regulatory domain, has two isoforms: β1 and β2 which are encoded from [[PRKAB1]] and [[PRKAB2]]
* γ, regulatory domain, has three isoforms: γ1, γ2, and γ3 which are encoded from [[PRKAG1]], [[PRKAG2]], and [[PRKAG3]]
In human skeletal muscle, the preferred form is α2β2γ1.<ref name=":2" /> But in the human liver, the most abundant form is α1β2γ1.<ref name=":2" />

== Mechanism ==
{{main|Alternative splicing}}
[[File:Alternative splicing.jpg|thumb|Different mechanisms of [[RNA splicing]]|320x320px]]
The primary mechanisms that produce protein isoforms are alternative splicing and variable promoter usage, though modifications due to genetic changes, such as [[Mutation|mutations]] and [[Polymorphism (biology)|polymorphisms]] are sometimes also considered distinct isoforms.<ref name=":5">{{cite journal | vauthors = Kornblihtt AR, Schor IE, Alló M, Dujardin G, Petrillo E, Muñoz MJ | title = Alternative splicing: a pivotal step between eukaryotic transcription and translation | language = En | journal = Nature Reviews Molecular Cell Biology | volume = 14 | issue = 3 | pages = 153–65 | date = March 2013 | pmid = 23385723 | doi = 10.1038/nrm3525 | s2cid = 54560052 | hdl = 11336/21049 | hdl-access = free }}</ref>

Alternative splicing is the main [[post-transcriptional modification]] process that produces mRNA transcript isoforms, and is a major molecular mechanism that may contribute to protein diversity.<ref name=":1" /> The [[spliceosome]], a large [[ribonucleoprotein]], is the molecular machine inside the nucleus responsible for RNA cleavage and [[Ligation (molecular biology)|ligation]], removing non-protein coding segments ([[Intron|introns]]).<ref name=":3">{{cite journal | vauthors = Lee Y, Rio DC | title = Mechanisms and Regulation of Alternative Pre-mRNA Splicing | journal = Annual Review of Biochemistry | volume = 84 | issue = 1 | pages = 291–323 | date = 2015-01-01 | pmid = 25784052 | pmc = 4526142 | doi = 10.1146/annurev-biochem-060614-034316 }}</ref>

Because splicing is a process that occurs between [[Transcription (biology)|transcription]] and [[Translation (biology)|translation]], its primary effects have mainly been studied through [[genomics]] techniques—for example, [[Microarray analysis techniques|microarray]] analyses and [[RNA-Seq|RNA sequencing]] have been used to identify alternatively spliced transcripts and measure their abundances.<ref name=":5" /> Transcript abundance is often used as a proxy for the abundance of protein isoforms, though [[proteomics]] experiments using gel electrophoresis and mass spectrometry have demonstrated that the correlation between transcript and protein counts is often low, and that one protein isoform is usually dominant.<ref name=":6">{{cite journal | vauthors = Tress ML, Abascal F, Valencia A | title = Alternative Splicing May Not Be the Key to Proteome Complexity | journal = Trends in Biochemical Sciences | volume = 42 | issue = 2 | pages = 98–110 | date = February 2017 | pmid = 27712956 | doi = 10.1016/j.tibs.2016.08.008 | pmc=6526280}}</ref> One 2015 study states that the cause of this discrepancy likely occurs after translation, though the mechanism is essentially unknown.<ref>{{cite journal | vauthors = Battle A, Khan Z, Wang SH, Mitrano A, Ford MJ, Pritchard JK, Gilad Y | title = Genomic variation. Impact of regulatory variation from RNA to protein | journal = Science | volume = 347 | issue = 6222 | pages = 664–7 | date = February 2015 | pmid = 25657249 | pmc = 4507520 | doi = 10.1126/science.1260793 }}</ref> Consequently, although alternative splicing has been implicated as an important link between variation and disease, there is no conclusive evidence that it acts primarily by producing novel protein isoforms.<ref name=":6" />

Alternative splicing generally describes a tightly regulated process in which alternative transcripts are intentionally generated by the splicing machinery. However, such transcripts are also produced by splicing errors in a process called "noisy splicing," and are also potentially translated into protein isoforms. Although ~95% of multi-exonic genes are thought to be alternatively spliced, one study on noisy splicing observed that most of the different low-abundance transcripts are noise, and predicts that most alternative transcript and protein isoforms present in a cell are not functionally relevant.<ref>{{cite journal | vauthors = Pickrell JK, Pai AA, Gilad Y, Pritchard JK | title = Noisy splicing drives mRNA isoform diversity in human cells | journal = PLOS Genetics | volume = 6 | issue = 12 | pages = e1001236 | date = December 2010 | pmid = 21151575 | pmc = 3000347 | doi = 10.1371/journal.pgen.1001236 | doi-access = free }}</ref>

Other transcriptional and post-transcriptional regulatory steps can also produce different protein isoforms.<ref>{{cite journal | vauthors = Smith LM, Kelleher NL | title = Proteoform: a single term describing protein complexity | language = En | journal = Nature Methods | volume = 10 | issue = 3 | pages = 186–7 | date = March 2013 | pmid = 23443629 | pmc = 4114032 | doi = 10.1038/nmeth.2369 }}</ref> Variable promoter usage occurs when the transcriptional machinery of a cell ([[RNA polymerase]], [[Transcription factor|transcription factors]], and other [[Enzyme|enzymes]]) begin transcription at different promoters—the region of DNA near a gene that serves as an initial binding site—resulting in slightly modified transcripts and protein isoforms.

== Characteristics ==
Generally, one protein isoform is labeled as the canonical sequence based on criteria such as its prevalence and similarity to [[Sequence homology|orthologous]]—or functionally analogous—sequences in other species.<ref>{{cite journal | vauthors = Li HD, Menon R, Omenn GS, Guan Y | title = Revisiting the identification of canonical splice isoforms through integration of functional genomics and proteomics evidence | journal = Proteomics | volume = 14 | issue = 23–24 | pages = 2709–18 | date = December 2014 | pmid = 25265570 | pmc = 4372202 | doi = 10.1002/pmic.201400170 | url = https://deepblue.lib.umich.edu/bitstream/2027.42/109787/1/pmic7911.pdf }}</ref> Isoforms are assumed to have similar functional properties, as most have similar sequences, and share some to most exons with the canonical sequence. However, some isoforms show much greater divergence (for example, through [[trans-splicing]]), and can share few to no exons with the canonical sequence. In addition, they can have different biological effects—for example, in an extreme case, the function of one isoform can promote cell survival, while another promotes cell death—or can have similar basic functions but differ in their sub-cellular localization.<ref>{{cite journal | vauthors = Sundvall M, Veikkolainen V, Kurppa K, Salah Z, Tvorogov D, van Zoelen EJ, Aqeilan R, Elenius K | title = Cell death or survival promoted by alternative isoforms of ErbB4 | journal = Molecular Biology of the Cell | volume = 21 | issue = 23 | pages = 4275–86 | date = December 2010 | pmid = 20943952 | pmc = 2993754 | doi = 10.1091/mbc.E10-04-0332 }}</ref> A 2016 study, however, functionally characterized all the isoforms of 1,492 genes and determined that most isoforms behave as "functional alloforms." The authors came to the conclusion that isoforms behave like distinct proteins after observing that the functional of most isoforms did not overlap.<ref>{{cite journal | vauthors = Yang X, Coulombe-Huntington J, Kang S, Sheynkman GM, Hao T, Richardson A, Sun S, Yang F, Shen YA, Murray RR, Spirohn K, Begg BE, Duran-Frigola M, MacWilliams A, Pevzner SJ, Zhong Q, Trigg SA, Tam S, Ghamsari L, Sahni N, Yi S, Rodriguez MD, Balcha D, Tan G, Costanzo M, Andrews B, Boone C, Zhou XJ, Salehi-Ashtiani K, Charloteaux B, Chen AA, Calderwood MA, Aloy P, Roth FP, Hill DE, Iakoucheva LM, Xia Y, Vidal M | display-authors = 6 | title = Widespread Expansion of Protein Interaction Capabilities by Alternative Splicing | journal = Cell | volume = 164 | issue = 4 | pages = 805–17 | date = February 2016 | pmid = 26871637 | pmc = 4882190 | doi = 10.1016/j.cell.2016.01.029 }}</ref> Because the study was conducted on cells ''in vitro'', it is not known if the isoforms in the expressed human proteome share these characteristics. Additionally, because the function of each isoform must generally be determined separately, most identified and predicted isoforms still have unknown functions.

==Related concepts==

=== Glycoform ===
{{main|Glycoprotein}}

A '''glycoform''' is an isoform of a protein that differs only with respect to the number or type of attached [[glycan]]. [[Glycoproteins]] often consist of a number of different glycoforms, with alterations in the attached [[saccharide]] or [[oligosaccharide]]. These modifications may result from differences in [[biosynthesis]] during the process of [[glycosylation]], or due to the action of [[glycosidases]] or [[glycosyltransferases]]. Glycoforms may be detected through detailed chemical analysis of separated glycoforms, but more conveniently detected through differential reaction with [[lectins]], as in [[lectin affinity chromatography]] and [[lectin]] [[affinity electrophoresis]]. Typical examples of glycoproteins consisting of glycoforms are the [[blood proteins]] as [[orosomucoid]], [[antitrypsin]], and [[haptoglobin]]. An unusual glycoform variation is seen in [[neural cell adhesion molecule|neuronal cell adhesion molecule, NCAM]] involving [[polysialic acid|polysialic acids, PSA]].

==Examples==

* [[G-actin]]: despite its conserved nature, it has a varying number of isoforms (at least six in mammals).
* [[Creatine kinase]], the presence of which in the blood can be used as an aid in the diagnosis of [[myocardial infarction]], exists in 3 isoforms.
* [[Hyaluronan synthase]], the enzyme responsible for the production of hyaluronan, has three isoforms in mammalian cells.
* [[UDP-glucuronosyltransferase]], an enzyme superfamily responsible for the detoxification pathway of many drugs, environmental pollutants, and toxic endogenous compounds has 16 known isoforms encoded in the human genome.<ref name="pmid17263731">{{cite journal | vauthors = Barre L, Fournel-Gigleux S, Finel M, Netter P, Magdalou J, Ouzzine M | title = Substrate specificity of the human UDP-glucuronosyltransferase UGT2B4 and UGT2B7. Identification of a critical aromatic amino acid residue at position 33 | journal = The FEBS Journal | volume = 274 | issue = 5 | pages = 1256–64 | date = March 2007 | pmid = 17263731 | doi = 10.1111/j.1742-4658.2007.05670.x | doi-access = free }}</ref>
*G6PDA: normal ratio of active isoforms in cells of any tissue is 1:1 shared with G6PDG. This is precisely the normal isoform ratio in hyperplasia. Only one of these isoforms is found during neoplasia.<ref>Pathoma, Fundamentals of Pathology</ref>
[[Monoamine oxidase]], a family of enzymes that catalyze the oxidation of monoamines, exists in two isoforms, MAO-A and MAO-B.

== See also ==
* [[Gene isoform]]

== References ==
{{Reflist|32em}}

== External links ==
{{Wiktionary|isoform}}
* [https://www.ncbi.nlm.nih.gov/entrez/query.fcgi?cmd=Retrieve&db=mesh&list_uids=68020033&dopt=Full MeSH entry protein isoforms]
* [http://ghr.nlm.nih.gov/glossary=isoforms Definitions Isoform]

{{Protein topics}}

{{DEFAULTSORT:Protein Isoform}}
[[Category:Protein structure]]