000119931 001__ 119931
000119931 005__ 20240319081018.0
000119931 0247_ $$2doi$$a10.1016/j.abb.2022.109392
000119931 0248_ $$2sideral$$a130771
000119931 037__ $$aART-2022-130771
000119931 041__ $$aeng
000119931 100__ $$aPacheco-Garcia, Juan Luis
000119931 245__ $$aDifferent phenotypic outcome due to site-specific phosphorylation in the cancer-associated NQO1 enzyme studied by phosphomimetic mutations
000119931 260__ $$c2022
000119931 5060_ $$aAccess copy available to the general public$$fUnrestricted
000119931 5203_ $$aProtein phosphorylation is a common phenomenon in human flavoproteins although the functional consequences of this site-specific modification are largely unknown. Here, we evaluated the effects of site-specific phosphorylation (using phosphomimetic mutations at sites S40, S82 and T128) on multiple functional aspects as well as in the structural stability of the antioxidant and disease-associated human flavoprotein NQO1 using biophysical and biochemical methods. In vitro biophysical studies revealed effects of phosphorylation at different sites such as decreased binding affinity for FAD and structural stability of its binding site (S82), conformational stability (S40 and S82) and reduced catalytic efficiency and functional cooperativity (T128). Local stability measurements by H/D exchange in different ligation states provided structural insight into these effects. Transfection of eukaryotic cells showed that phosphorylation at sites S40 and S82 may reduce steady-levels of NQO1 protein by enhanced proteasome-induced degradation. We show that site-specific phosphorylation of human NQO1 may cause pleiotropic and counterintuitive effects on this multifunctional protein with potential implications for its relationships with human disease. Our approach allows to establish relationships between site-specific phosphorylation, functional and structural stability effects in vitro and inside cells paving the way for more detailed analyses of phosphorylation at the flavoproteome scale.
000119931 536__ $$9info:eu-repo/grantAgreement/ES/DGA-FEDER/E35-20R$$9info:eu-repo/grantAgreement/ES/MCIU-ERDF/RTI2018-096246-B-I00$$9info:eu-repo/grantAgreement/ES/MICINN-AEI/PID2019-103901GB-I00
000119931 540__ $$9info:eu-repo/semantics/openAccess$$aby-nc-nd$$uhttp://creativecommons.org/licenses/by-nc-nd/3.0/es/
000119931 590__ $$a3.9$$b2022
000119931 592__ $$a0.867$$b2022
000119931 591__ $$aBIOPHYSICS$$b17 / 70 = 0.243$$c2022$$dQ1$$eT1
000119931 593__ $$aBiophysics$$c2022$$dQ1
000119931 591__ $$aBIOCHEMISTRY & MOLECULAR BIOLOGY$$b126 / 285 = 0.442$$c2022$$dQ2$$eT2
000119931 593__ $$aMolecular Biology$$c2022$$dQ2
000119931 593__ $$aBiochemistry$$c2022$$dQ2
000119931 594__ $$a7.5$$b2022
000119931 655_4 $$ainfo:eu-repo/semantics/article$$vinfo:eu-repo/semantics/publishedVersion
000119931 700__ $$aAnoz Carbonell, Ernesto
000119931 700__ $$aLoginov, Dmitry S.
000119931 700__ $$aVankova, Pavla
000119931 700__ $$aSalido, Eduardo
000119931 700__ $$aMan, Petr
000119931 700__ $$0(orcid)0000-0001-8743-0182$$aMedina, Milagros$$uUniversidad de Zaragoza
000119931 700__ $$aPalomino-Morales, Rogelio
000119931 700__ $$aPey, Angel L.
000119931 7102_ $$11002$$2060$$aUniversidad de Zaragoza$$bDpto. Bioq.Biolog.Mol. Celular$$cÁrea Bioquímica y Biolog.Mole.
000119931 773__ $$g729 (2022), 109392 [12 pp]$$pArch. biochem. biophys.$$tARCHIVES OF BIOCHEMISTRY AND BIOPHYSICS$$x0003-9861
000119931 8564_ $$s6086056$$uhttps://zaguan.unizar.es/record/119931/files/texto_completo.pdf$$yVersión publicada
000119931 8564_ $$s2642627$$uhttps://zaguan.unizar.es/record/119931/files/texto_completo.jpg?subformat=icon$$xicon$$yVersión publicada
000119931 909CO $$ooai:zaguan.unizar.es:119931$$particulos$$pdriver
000119931 951__ $$a2024-03-18-15:53:02
000119931 980__ $$aARTICLE