Editing Pyruvate kinase (section)

==Regulation==
Glycolysis is highly regulated at three of its catalytic steps: the phosphorylation of glucose by [[hexokinase]], the phosphorylation of [[Fructose 6-phosphate|fructose-6-phosphate]] by [[phosphofructokinase]], and the transfer of phosphate from PEP to ADP by pyruvate kinase. Under wild-type conditions, all three of these reactions are irreversible, have a large negative free energy and are responsible for the regulation of this pathway.<ref name="Berg_2002" /> Pyruvate kinase activity is most broadly regulated by allosteric effectors, covalent modifiers and hormonal control. However, the most significant pyruvate kinase regulator is [[Fructose 1,6-bisphosphate|fructose-1,6-bisphosphate]] (FBP), which serves as an allosteric effector for the enzyme.

=== Allosteric effectors ===
[[Allosteric regulation]] is the binding of an effector to a site on the protein other than the active site, causing a conformational change and altering the activity of that given protein or enzyme. Pyruvate kinase has been found to be allosterically activated by FBP and allosterically inactivated by ATP and alanine.<ref>{{cite journal | vauthors = Carbonell J, Felíu JE, Marco R, Sols A | title = Pyruvate kinase. Classes of regulatory isoenzymes in mammalian tissues | journal = European Journal of Biochemistry | volume = 37 | issue = 1 | pages = 148–56 | date = August 1973 | pmid = 4729424 | doi = 10.1111/j.1432-1033.1973.tb02969.x | hdl-access = free | hdl = 10261/78345 }}</ref> Pyruvate Kinase tetramerization is promoted by FBP and Serine while tetramer dissociation is promoted by L-Cysteine.<ref>{{cite journal | vauthors = Yang J, Liu H, Liu X, Gu C, Luo R, Chen HF | title = Synergistic Allosteric Mechanism of Fructose-1,6-bisphosphate and Serine for Pyruvate Kinase M2 via Dynamics Fluctuation Network Analysis | journal = Journal of Chemical Information and Modeling | volume = 56 | issue = 6 | pages = 1184–1192 | date = June 2016 | pmid = 27227511 | pmc = 5115163 | doi = 10.1021/acs.jcim.6b00115 }}</ref><ref>{{cite journal | vauthors = Chaneton B, Hillmann P, Zheng L, Martin AC, Maddocks OD, Chokkathukalam A, Coyle JE, Jankevics A, Holding FP, Vousden KH, Frezza C, O'Reilly M, Gottlieb E | display-authors = 6 | title = Serine is a natural ligand and allosteric activator of pyruvate kinase M2 | journal = Nature | volume = 491 | issue = 7424 | pages = 458–462 | date = November 2012 | pmid = 23064226 | pmc = 3894725 | doi = 10.1038/nature11540 | bibcode = 2012Natur.491..458C }}</ref><ref>{{cite journal | vauthors = Nakatsu D, Horiuchi Y, Kano F, Noguchi Y, Sugawara T, Takamoto I, Kubota N, Kadowaki T, Murata M | display-authors = 6 | title = L-cysteine reversibly inhibits glucose-induced biphasic insulin secretion and ATP production by inactivating PKM2 | journal = Proceedings of the National Academy of Sciences of the United States of America | volume = 112 | issue = 10 | pages = E1067-76 | date = March 2015 | pmid = 25713368 | pmc = 4364213 | doi = 10.1073/pnas.1417197112 | bibcode = 2015PNAS..112E1067N | doi-access = free }}</ref>

==== Fructose-1,6-bisphosphate ====
FBP is the most significant source of regulation because it comes from within the glycolysis pathway. FBP is a glycolytic intermediate produced from the phosphorylation of [[fructose 6-phosphate]]. FBP binds to the allosteric binding site on domain C of pyruvate kinase and changes the conformation of the enzyme, causing the activation of pyruvate kinase activity.<ref>{{cite journal | vauthors = Ishwar A |title=Distinguishing the interactions in the fructose 1,6-bisphosphate binding site of human liver pyruvate kinase that contribute to allostery. |journal=Biochemistry |volume=54 |issue=7 |pages=1516–24 |date=24 February 2015 |pmid=25629396 |pmc=5286843 |doi=10.1021/bi501426w }}</ref> As an intermediate present within the glycolytic pathway, FBP provides [[feed forward (control)|feedforward stimulation]] because the higher the concentration of FBP, the greater the allosteric activation and magnitude of pyruvate kinase activity. Pyruvate kinase is most sensitive to the effects of FBP. As a result, the remainder of the regulatory mechanisms serve as secondary modification.<ref name="Valentini 18145–18152"/><ref>{{cite journal | vauthors = Jurica MS, Mesecar A, Heath PJ, Shi W, Nowak T, Stoddard BL | title = The allosteric regulation of pyruvate kinase by fructose-1,6-bisphosphate | journal = Structure | volume = 6 | issue = 2 | pages = 195–210 | date = February 1998 | pmid = 9519410 | doi = 10.1016/S0969-2126(98)00021-5 | doi-access = free }}</ref>

=== Covalent modifiers ===
Covalent modifiers serve as indirect regulators by controlling the phosphorylation, dephosphorylation, acetylation, succinylation and oxidation of enzymes, resulting in the activation and inhibition of enzymatic activity.<ref>{{cite journal | vauthors = Li YH, Li XF, Liu JT, Wang H, Fan LL, Li J, Sun GP | title = PKM2, a potential target for regulating cancer | journal = Gene | volume = 668 | pages = 48–53 | date = August 2018 | pmid = 29775756 | doi = 10.1016/j.gene.2018.05.038 | s2cid = 205030574 }}</ref> In the liver, [[glucagon]] and [[epinephrine]] activate [[protein kinase A]], which serves as a covalent modifier by phosphorylating and deactivating pyruvate kinase. In contrast, the secretion of insulin in response to blood sugar elevation activates phosphoprotein phosphatase I, causing the dephosphorylation and activation of pyruvate kinase to increase glycolysis. The same covalent modification has the opposite effect on gluconeogenesis enzymes. This regulation system is responsible for the avoidance of a futile cycle through the prevention of simultaneous activation of pyruvate kinase and enzymes that catalyze gluconeogenesis.<ref>{{cite journal | vauthors = Birnbaum MJ, Fain JN | title = Activation of protein kinase and glycogen phosphorylase in isolated rat liver cells by glucagon and catecholamines | journal = The Journal of Biological Chemistry | volume = 252 | issue = 2 | pages = 528–35 | date = January 1977 | doi = 10.1016/S0021-9258(17)32749-7 | pmid = 188818 | doi-access = free }}</ref>

=== Hormonal control ===
In order to prevent a [[futile cycle]], glycolysis and gluconeogenesis are heavily regulated in order to ensure that they are never operating in the cell at the same time. As a result, the inhibition of pyruvate kinase by glucagon, cyclic AMP and epinephrine, not only shuts down glycolysis, but also stimulates gluconeogenesis. Alternatively, insulin interferes with the effect of glucagon, cyclic AMP and epinephrine, causing pyruvate kinase to function normally and gluconeogenesis to be shut down. Furthermore, glucose was found to inhibit and disrupt gluconeogenesis, leaving pyruvate kinase activity and glycolysis unaffected. Overall, the interaction between hormones plays a key role in the functioning and regulation of glycolysis and gluconeogenesis in the cell.<ref name="Feliú_1976">{{cite journal | vauthors = Feliú JE, Hue L, Hers HG | title = Hormonal control of pyruvate kinase activity and of gluconeogenesis in isolated hepatocytes | journal = Proceedings of the National Academy of Sciences of the United States of America | volume = 73 | issue = 8 | pages = 2762–6 | year = 1976 | pmid = 183209 | pmc = 430732 | doi = 10.1073/pnas.73.8.2762| bibcode = 1976PNAS...73.2762F | doi-access = free }}</ref>

==== Inhibitory effect of metformin ====
Metformin, or [[Metformin|dimethylbiguanide]], is the primary treatment used for type 2 diabetes. Metformin has been shown to indirectly affect pyruvate kinase through the inhibition of gluconeogenesis. Specifically, the addition of metformin is linked to a marked decrease in glucose flux and increase in lactate/pyruvate flux from various metabolic pathways. Although metformin does not directly affect pyruvate kinase activity, it causes a decrease in the concentration of ATP. Due to the allosteric inhibitory effects of ATP on pyruvate kinase, a decrease in ATP results in diminished inhibition and the subsequent stimulation of pyruvate kinase. Consequently, the increase in pyruvate kinase activity directs metabolic flux through glycolysis rather than gluconeogenesis.<ref name="Argaud_1993">{{cite journal | vauthors = Argaud D, Roth H, Wiernsperger N, Leverve XM | title = Metformin decreases gluconeogenesis by enhancing the pyruvate kinase flux in isolated rat hepatocytes | journal = European Journal of Biochemistry | volume = 213 | issue = 3 | pages = 1341–8 | year = 1993 | pmid = 8504825 | doi = 10.1111/j.1432-1033.1993.tb17886.x | doi-access = free }}</ref>

=== Gene Regulation ===
[[Heterogeneous ribonucleoprotein particle|Heterogenous ribonucleotide proteins]] (hnRNPs) can act on the PKM gene to regulate expression of M1 and M2 isoforms. PKM1 and PKM2 isoforms are splice variants of the PKM gene that differ by a single exon. Various types of hnRNPs such as hnRNPA1 and hnRNPA2 enter the nucleus during hypoxia conditions and modulate expression such that PKM2 is up-regulated.<ref>{{cite journal | vauthors = Clower CV, Chatterjee D, Wang Z, Cantley LC, Vander Heiden MG, Krainer AR | title = The alternative splicing repressors hnRNP A1/A2 and PTB influence pyruvate kinase isoform expression and cell metabolism | journal = Proceedings of the National Academy of Sciences of the United States of America | volume = 107 | issue = 5 | pages = 1894–9 | date = February 2010 | pmid = 20133837 | pmc = 2838216 | doi = 10.1073/pnas.0914845107 | bibcode = 2010PNAS..107.1894C | doi-access = free }}</ref> Hormones such as [[insulin]] up-regulate expression of PKM2 while hormones like [[Triiodothyronine|tri-iodothyronine]] (T3) and [[glucagon]] aid in down-regulating PKM2.<ref>{{cite journal | vauthors = Iqbal MA, Siddiqui FA, Gupta V, Chattopadhyay S, Gopinath P, Kumar B, Manvati S, Chaman N, Bamezai RN | display-authors = 6 | title = Insulin enhances metabolic capacities of cancer cells by dual regulation of glycolytic enzyme pyruvate kinase M2 | journal = Molecular Cancer | volume = 12 | issue = 1 | pages = 72 | date = July 2013 | pmid = 23837608 | pmc = 3710280 | doi = 10.1186/1476-4598-12-72 | doi-access = free }}</ref>

==== Carbohydrate response element binding protein (ChREBP) ====
[[Carbohydrate-responsive element-binding protein|ChREBP]] is a [[transcription factor]] that regulates expression of the L isozyme of pyruvate kinase.<ref name=":0">{{cite journal |vauthors=Kawaguchi T, Takenoshita M, Kabashima T, Uyeda K |date=November 2001 |title=Glucose and cAMP regulate the L-type pyruvate kinase gene by phosphorylation/dephosphorylation of the carbohydrate response element binding protein |journal=Proceedings of the National Academy of Sciences of the United States of America |volume=98 |issue=24 |pages=13710–5 |bibcode=2001PNAS...9813710K |doi=10.1073/pnas.231370798 |pmc=61106 |pmid=11698644 |doi-access=free}}</ref> A glucose-sensing module contains domains that are targets for regulatory phosphorylation based on the concentrations of glucose and cAMP, which then control its import into the nucleus.<ref name=":1">{{Cite journal |last1=Ortega-Prieto |first1=Paula |last2=Postic |first2=Catherine |date=2019 |title=Carbohydrate Sensing Through the Transcription Factor ChREBP |journal=Frontiers in Genetics |volume=10 |page=472 |doi=10.3389/fgene.2019.00472 |pmid=31275349 |pmc=6593282 |issn=1664-8021|doi-access=free }}</ref> It may also be further activated by directly binding [[Glucose 6-phosphate|glucose-6-phosphate.]]<ref name=":0" /><ref>{{Cite journal |last1=Richards |first1=Paul |last2=Ourabah |first2=Sarah |last3=Montagne |first3=Jacques |last4=Burnol |first4=Anne-Françoise |last5=Postic |first5=Catherine |last6=Guilmeau |first6=Sandra |date=2017 |title=MondoA/ChREBP: The usual suspects of transcriptional glucose sensing; Implication in pathophysiology |url=https://pubmed.ncbi.nlm.nih.gov/28403938 |journal=Metabolism: Clinical and Experimental |volume=70 |pages=133–151 |doi=10.1016/j.metabol.2017.01.033 |issn=1532-8600 |pmid=28403938}}</ref> Once in the nucleus, its DNA binding domains activate pyruvate kinase transcription.<ref name=":1" /> Therefore, high glucose and low cAMP causes dephosphorylation of [[Carbohydrate-responsive element-binding protein|ChREBP]], which then upregulates expression of pyruvate kinase in the liver.<ref name=":0" />