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In the methanogenic archae Methanosarcina barkeri, insertion of pyrrolysine, the 22nd amino acid, results from the decoding of an amber UAG codon in the mRNA of monomethylamine methyltransferases (MtmB). Sequence comparisons combined with structural enzymatic and chemical probing on M. barkeri MtmB1 mRNA demonstrate the presence of a hairpin motif located immediately after the redefined UAG codon. This structure of 86 nucleotides differs slightly from a proposal given in the literature and comprises four successive stems separated by three internal loops and closed by a large apical loop. Sequence alignments of MtmB mRNAs of different Methanosarcinacae reveal a conservation of the motif in both sequence and folding levels. The functional role of this motif as a signal leading to pyrrolysine insertion is discussed.
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http://dx.doi.org/10.1016/j.biochi.2005.03.006 | DOI Listing |
J Bacteriol
February 2023
BGI Research-Shenzhen, BGI, Shenzhen, China.
The cotranslational incorporation of pyrrolysine (Pyl), the 22nd proteinogenic amino acid, into proteins in response to the UAG stop codon represents an outstanding example of natural genetic code expansion. Genetic encoding of Pyl is conducted by the pyrrolysyl-tRNA synthetase (PylRS) and its cognate tRNA, tRNA. Owing to the high tolerance of PylRS toward diverse amino acid substrates and great orthogonality in various model organisms, the PylRS/tRNA-derived pairs are ideal for genetic code expansion to insert noncanonical amino acids (ncAAs) into proteins of interest.
View Article and Find Full Text PDFBiosystems
October 2020
Aix-Marseille University, IRD, VITROME, Institut Hospitalo-Universitaire Méditerranée-Infection, Marseille, France. Electronic address:
During the noncanonical deletion transcription, k nucleotides are systematically skipped/deleted after each transcribed trinucleotide producing deletion-RNAs (delRNAs). Peptides matching delRNAs either result from (a) canonical translation of delRNAs; or (b) noncanonical translation of regular transcripts along expanded codons. Only along frame "0" (start site) (a) and (b) produce identical peptides.
View Article and Find Full Text PDFLife (Basel)
February 2017
Division of Life Science and Applied Genomics Center, Hong Kong University of Science & Technology, Clear Water Bay, Hong Kong, China.
The methods for establishing synthetic lifeforms with rewritten genetic codes comprising non-canonical amino acids (NCAA) in addition to canonical amino acids (CAA) include proteome-wide replacement of CAA, insertion through suppression of nonsense codon, and insertion via the pyrrolysine and selenocysteine pathways. Proteome-wide reassignments of nonsense codons and sense codons are also under development. These methods enable the application of NCAAs to enrich both fundamental and applied aspects of protein chemistry and biology.
View Article and Find Full Text PDFCroat Chem Acta
June 2016
Department of Molecular Biophysics & Biochemistry, Yale University, New Haven, CT 06520-8114, USA.
Genetic code expansion (GCE) has become a central topic of synthetic biology. GCE relies on engineered aminoacyl-tRNA synthetases (aaRSs) and a cognate tRNA species to allow codon reassignment by co-translational insertion of non-canonical amino acids (ncAAs) into proteins. Introduction of such amino acids increases the chemical diversity of recombinant proteins endowing them with novel properties.
View Article and Find Full Text PDFJ Am Chem Soc
March 2015
∥Departments of Biochemistry and Chemistry, Duke University School of Medicine, Durham, North Carolina 27710, United States.
Mammalian mitochondrial tRNA(Ser(UCN)) (mt-tRNA(Ser)) and pyrrolysine tRNA (tRNA(Pyl)) fold to near-canonical three-dimensional structures despite having noncanonical secondary structures with shortened interhelical loops that disrupt the conserved tRNA tertiary interaction network. How these noncanonical tRNAs compensate for their loss of tertiary interactions remains unclear. Furthermore, in human mt-tRNA(Ser), lengthening the variable loop by the 7472insC mutation reduces mt-tRNA(Ser) concentration in vivo through poorly understood mechanisms and is strongly associated with diseases such as deafness and epilepsy.
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