000130479 001__ 130479
000130479 005__ 20240125162931.0
000130479 0247_ $$2doi$$a10.1074/jbc.RA120.015103
000130479 0248_ $$2sideral$$a122576
000130479 037__ $$aART-2021-122576
000130479 041__ $$aeng
000130479 100__ $$0(orcid)0000-0002-6794-3535$$aLatorre-Muro, Pedro
000130479 245__ $$aSelf-acetylation at the active site of phosphoenolpyruvate carboxykinase (PCK1) controls enzyme activity
000130479 260__ $$c2021
000130479 5060_ $$aAccess copy available to the general public$$fUnrestricted
000130479 5203_ $$aAcetylation is known to regulate the activity of cytosolicphosphoenolpyruvate carboxykinase (PCK1), a key enzyme ingluconeogenesis, by promoting the reverse reaction of theenzyme (converting phosphoenolpyruvate to oxaloacetate). It isalso known that the histone acetyltransferase p300 can inducePCK1 acetylation in cells, but whether that is a direct or in-direct function was not known. Here we initially set out todetermine whether p300 can acetylate directly PCK1in vitro.We report that p300 weakly acetylates PCK1, but surprisingly,using several techniques including protein crystallization, massspectrometry, isothermal titration calorimetry, saturation-transfer difference nuclear magnetic resonance and moleculardocking, we found that PCK1 is also able to acetylate itselfusing acetyl-CoA independently of p300. This reaction yieldedan acetylated recombinant PCK1 with a 3-fold decrease inkcatwithout changes inKmfor all substrates. Acetylation stoichi-ometry was determined for 14 residues, including residueslining the active site. Structural and kinetic analyses deter-mined that site-directed acetylation of K244, located inside theactive site, altered this site and rendered the enzyme inactive.In addition, we found that acetyl-CoA binding to the active siteis specific and metal dependent. Ourfindings provide directevidence for acetyl-CoA binding and chemical reaction withthe active site of PCK1 and suggest a newly discovered regu-latory mechanism of PCK1 during metabolic stress.
000130479 536__ $$9info:eu-repo/grantAgreement/ES/DGA/E45-17R$$9info:eu-repo/grantAgreement/ES/MEC/AGL2015–66177-R$$9info:eu-repo/grantAgreement/ES/UZ/2015-BIO-01
000130479 540__ $$9info:eu-repo/semantics/openAccess$$aby$$uhttp://creativecommons.org/licenses/by/3.0/es/
000130479 590__ $$a5.485$$b2021
000130479 591__ $$aBIOCHEMISTRY & MOLECULAR BIOLOGY$$b94 / 297 = 0.316$$c2021$$dQ2$$eT1
000130479 592__ $$a1.871$$b2021
000130479 593__ $$aCell Biology$$c2021$$dQ1
000130479 593__ $$aBiochemistry$$c2021$$dQ1
000130479 594__ $$a8.8$$b2021
000130479 655_4 $$ainfo:eu-repo/semantics/article$$vinfo:eu-repo/semantics/publishedVersion
000130479 700__ $$aBaeza, Josué
000130479 700__ $$0(orcid)0000-0002-3122-9401$$aHurtado-Guerrero, Ramón
000130479 700__ $$aHicks, Thomas
000130479 700__ $$0(orcid)0000-0001-8355-2289$$aDelso, Ignacio$$uUniversidad de Zaragoza
000130479 700__ $$aHernández-Ruiz, Cristina
000130479 700__ $$0(orcid)0000-0001-5702-4538$$aVelazquez-Campoy, Adrián$$uUniversidad de Zaragoza
000130479 700__ $$aLawton, Alexis J.
000130479 700__ $$aAngulo, Jesús
000130479 700__ $$aDenu, John M.
000130479 700__ $$0(orcid)0000-0003-0062-1029$$aCarrodeguas, José A.$$uUniversidad de Zaragoza
000130479 7102_ $$12013$$2765$$aUniversidad de Zaragoza$$bDpto. Química Orgánica$$cÁrea Química Orgánica
000130479 7102_ $$11002$$2060$$aUniversidad de Zaragoza$$bDpto. Bioq.Biolog.Mol. Celular$$cÁrea Bioquímica y Biolog.Mole.
000130479 773__ $$g296 (2021), 100205 [13 pp.]$$pJ. biol. chem.$$tJournal of Biological Chemistry$$x0021-9258
000130479 8564_ $$s3228615$$uhttps://zaguan.unizar.es/record/130479/files/texto_completo.pdf$$yVersión publicada
000130479 8564_ $$s3602897$$uhttps://zaguan.unizar.es/record/130479/files/texto_completo.jpg?subformat=icon$$xicon$$yVersión publicada
000130479 909CO $$ooai:zaguan.unizar.es:130479$$particulos$$pdriver
000130479 951__ $$a2024-01-25-15:14:01
000130479 980__ $$aARTICLE