000048440 001__ 48440
000048440 005__ 20161006102705.0
000048440 0247_ $$2doi$$a10.1038/srep09129
000048440 0248_ $$2sideral$$a93811
000048440 037__ $$aART-2015-93811
000048440 041__ $$aeng
000048440 100__ $$aLlamazares, Emilio
000048440 245__ $$aRational stabilization of complex proteins: A divide and combine approach
000048440 260__ $$c2015
000048440 5060_ $$aAccess copy available to the general public$$fUnrestricted
000048440 5203_ $$aIncreasing the thermostability of proteins is often crucial for their successful use as analytic, synthetic or therapeutic tools. Most rational thermostabilization strategies were developed on small two-state proteins and, unsurprisingly, they tend to fail when applied to the much more abundant, larger, non-fully cooperative proteins. We show that the key to stabilize the latter is to know the regions of lower stability. To prove it, we have engineered apoflavodoxin, a non-fully cooperative protein on which previous thermostabilizing attempts had failed. We use a step-wise combination of structure-based, rationally-designed, stabilizing mutations confined to the less stable structural region, and obtain variants that, according to their van't Hoff to calorimetric enthalpy ratios, exhibit fully-cooperative thermal unfolding with a melting temperature of 75°C, 32 degrees above the lower melting temperature of the non-cooperative wild type protein. The ideas introduced here may also be useful for the thermostabilization of complex proteins through formulation or using specific stabilizing ligands (e.g. pharmacological chaperones).
000048440 536__ $$9info:eu-repo/grantAgreement/ES/DGA/B89$$9info:eu-repo/grantAgreement/ES/MICINN/BFU2010-16297$$9info:eu-repo/grantAgreement/ES/MINECO/BFU2013-47064-P
000048440 540__ $$9info:eu-repo/semantics/openAccess$$aby$$uhttp://creativecommons.org/licenses/by/3.0/es/
000048440 590__ $$a5.228$$b2015
000048440 591__ $$aMULTIDISCIPLINARY SCIENCES$$b7 / 60 = 0.117$$c2015$$dQ1$$eT1
000048440 655_4 $$ainfo:eu-repo/semantics/article$$vinfo:eu-repo/semantics/publishedVersion
000048440 700__ $$0(orcid)0000-0003-4597-4020$$aClemente Arellano, Isabel$$uUniversidad de Zaragoza
000048440 700__ $$aBueno Fernandez, Marta
000048440 700__ $$0(orcid)0000-0001-5702-4538$$aVelázquez Campoy, Adrián$$uUniversidad de Zaragoza
000048440 700__ $$0(orcid)0000-0002-2879-9200$$aSancho Sanz, Javier$$uUniversidad de Zaragoza
000048440 7102_ $$12009$$2750$$aUniversidad de Zaragoza$$bDepartamento de Química Analítica$$cQuímica Analítica
000048440 7102_ $$11002$$2060$$aUniversidad de Zaragoza$$bDepartamento de Bioquímica y Biología Molecular y Celular$$cBioquímica y Biología Molecular
000048440 773__ $$g5 (2015), 9129 [11 pp.]$$pSci. rep.$$tSCIENTIFIC REPORTS$$x2045-2322
000048440 8564_ $$s1984482$$uhttp://zaguan.unizar.es/record/48440/files/texto_completo.pdf$$yVersión publicada
000048440 8564_ $$s102771$$uhttp://zaguan.unizar.es/record/48440/files/texto_completo.jpg?subformat=icon$$xicon$$yVersión publicada
000048440 909CO $$ooai:zaguan.unizar.es:48440$$particulos$$pdriver
000048440 951__ $$a2016-06-27-11:21:22
000048440 980__ $$aARTICLE