Engineering Pyrrolysyl-tRNA Synthetase for the Incorporation of Non-Canonical Amino Acids with Smaller Side Chains

dc.contributor.authorKoch, Nikolaj G.
dc.contributor.authorGoettig, Peter
dc.contributor.authorRappsilber, Juri
dc.contributor.authorBudisa, Nediljko
dc.date.accessioned2022-09-12T14:09:44Z
dc.date.available2022-09-12T14:09:44Z
dc.date.issued2021-10-17
dc.date.updated2022-09-03T10:32:17Z
dc.description.abstractSite-specific incorporation of non-canonical amino acids (ncAAs) into proteins has emerged as a universal tool for systems bioengineering at the interface of chemistry, biology, and technology. The diversification of the repertoire of the genetic code has been achieved for amino acids with long and/or bulky side chains equipped with various bioorthogonal tags and useful spectral probes. Although ncAAs with relatively small side chains and similar properties are of great interest to biophysics, cell biology, and biomaterial science, they can rarely be incorporated into proteins. To address this gap, we report the engineering of PylRS variants capable of incorporating an entire library of aliphatic “small-tag” ncAAs. In particular, we performed mutational studies of a specific PylRS, designed to incorporate the shortest non-bulky ncAA (S-allyl-l-cysteine) possible to date and based on this knowledge incorporated aliphatic ncAA derivatives. In this way, we have not only increased the number of translationally active “small-tag” ncAAs, but also determined key residues responsible for maintaining orthogonality, while engineering the PylRS for these interesting substrates. Based on the known plasticity of PylRS toward different substrates, our approach further expands the reassignment capacities of this enzyme toward aliphatic amino acids with smaller side chains endowed with valuable functionalities.en
dc.description.sponsorshipBMBF, 031B0584A, IBÖM04: XenoGlue - Ein neuartiger Muschel-basierter fotoaktivierbarer Bioklebstoff für biomedizinische Anwendungenen
dc.description.sponsorshipDFG, 390540038, EXC 2008: Unifying Systems in Catalysis "UniSysCat"en
dc.identifier.eissn1422-0067
dc.identifier.issn1661-6596
dc.identifier.urihttps://depositonce.tu-berlin.de/handle/11303/17433
dc.identifier.urihttp://dx.doi.org/10.14279/depositonce-16214
dc.language.isoenen
dc.rights.urihttps://creativecommons.org/licenses/by/4.0/en
dc.subject.ddc540 Chemiede
dc.subject.ddc570 Biowissenschaften; Biologiede
dc.subject.othergenetic code expansionen
dc.subject.otherpyrrolysyl-tRNA synthetasesen
dc.subject.othernon-canonical amino acidsen
dc.subject.otheraliphatic amino acidsen
dc.subject.otherbioorthogonal reactive handlesen
dc.subject.otherazidohomoalanineen
dc.subject.otherphoto-methionineen
dc.subject.otherprotein engineeringen
dc.subject.otherstop codon suppressionen
dc.subject.otherS-allyl-l-cysteineen
dc.titleEngineering Pyrrolysyl-tRNA Synthetase for the Incorporation of Non-Canonical Amino Acids with Smaller Side Chainsen
dc.typeArticleen
dc.type.versionpublishedVersionen
dcterms.bibliographicCitation.articlenumber11194en
dcterms.bibliographicCitation.doi10.3390/ijms222011194en
dcterms.bibliographicCitation.issue20en
dcterms.bibliographicCitation.journaltitleInternational Journal of Molecular Sciencesen
dcterms.bibliographicCitation.originalpublishernameMDPIen
dcterms.bibliographicCitation.originalpublisherplaceBaselen
dcterms.bibliographicCitation.volume22en
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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