Computational Aminoacyl-tRNA Synthetase Library Design for Photocaged Tyrosine

dc.contributor.authorBaumann, Tobias
dc.contributor.authorHauf, Matthias
dc.contributor.authorRichter, Florian
dc.contributor.authorAlbers, Suki
dc.contributor.authorMöglich, Andreas
dc.contributor.authorIgnatova, Zoya
dc.contributor.authorBudisa, Nediljko
dc.date.accessioned2019-05-14T08:37:03Z
dc.date.available2019-05-14T08:37:03Z
dc.date.issued2019-05-11
dc.description.abstractEngineering aminoacyl-tRNA synthetases (aaRSs) provides access to the ribosomal incorporation of noncanonical amino acids via genetic code expansion. Conventional targeted mutagenesis libraries with 5–7 positions randomized cover only marginal fractions of the vast sequence space formed by up to 30 active site residues. This frequently results in selection of weakly active enzymes. To overcome this limitation, we use computational enzyme design to generate a focused library of aaRS variants. For aaRS enzyme redesign, photocaged ortho-nitrobenzyl tyrosine (ONBY) was chosen as substrate due to commercial availability and its diverse applications. Diversifying 17 first- and second-shell sites and performing conventional aaRS positive and negative selection resulted in a high-activity aaRS. This MjTyrRS variant carries ten mutations and outperforms previously reported ONBY-specific aaRS variants isolated from traditional libraries. In response to a single in-frame amber stop codon, it mediates the in vivo incorporation of ONBY with an efficiency matching that of the wild type MjTyrRS enzyme acylating cognate tyrosine. These results exemplify an improved general strategy for aaRS library design and engineering.en
dc.description.sponsorshipDFG, 414044773, Open Access Publizieren 2019 - 2020 / Technische Universität Berlinen
dc.description.sponsorshipDFG, 207100805, FOR 1805: Einfluss der Ribosomendynamik auf Regulation der Geschwindigkeit und Genauigkeit der Translationen
dc.description.sponsorshipDFG, 390540038, EXC 2008: Vereinigung von Systemen in der Katalyseen
dc.identifier.eissn1422-0067
dc.identifier.urihttps://depositonce.tu-berlin.de/handle/11303/9428
dc.identifier.urihttp://dx.doi.org/10.14279/depositonce-8486
dc.language.isoenen
dc.rights.urihttps://creativecommons.org/licenses/by/4.0/en
dc.subject.ddc570 Biowissenschaften; Biologiede
dc.subject.ddc540 Chemie und zugeordnete Wissenschaftende
dc.subject.otherenzyme designen
dc.subject.othernoncanonical amino acidsen
dc.subject.otherprotein modificationen
dc.subject.otherdirected evolutionen
dc.subject.othermutagenesisen
dc.subject.othergene librariesen
dc.subject.othergenetic code expansionen
dc.subject.otherunnatural amino acidsen
dc.subject.otherprotein engineeringen
dc.titleComputational Aminoacyl-tRNA Synthetase Library Design for Photocaged Tyrosineen
dc.typeArticleen
dc.type.versionpublishedVersionen
dcterms.bibliographicCitation.articlenumber2343en
dcterms.bibliographicCitation.doi10.3390/ijms20092343en
dcterms.bibliographicCitation.issue9en
dcterms.bibliographicCitation.journaltitleInternational Journal of Molecular Sciencesen
dcterms.bibliographicCitation.originalpublishernameMDPIen
dcterms.bibliographicCitation.originalpublisherplaceBaselen
dcterms.bibliographicCitation.volume20en
tub.accessrights.dnbfreeen
tub.affiliationFak. 2 Mathematik und Naturwissenschaften::Inst. Chemie::FG Biokatalysede
tub.affiliation.facultyFak. 2 Mathematik und Naturwissenschaftende
tub.affiliation.groupFG Biokatalysede
tub.affiliation.instituteInst. Chemiede
tub.publisher.universityorinstitutionTechnische Universität Berlinen

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