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The expanding world of DNA and RNA

T. Chen, N. Hongdilokkul, Z. Liu, D. Thirunavukarasu, F.E. Romesberg, Curr. Op. Chem. Biol. (2016) 34:80–87.
pubpic2016bchenWe review nucleotide modifications, such as those to phosphate and sugar moieties that increase nuclease resistance or the range of activities possible, as well as whole nucleobase replacement that results in selective pairing and the creation of unnatural base pairs. Both in vitro and in vivo examples are discussed, including efforts to create semi-synthetic organisms with altered or expanded genetic alphabets.

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Evolution of thermophilic DNA polymerases for the recognition and amplification of C2′-modified DNA

T. Chen, N. Hongdilokkul, Z. Liu, R. Adhikary, S.S. Tsuen, F.E. Romesberg, Nat. Chem. (2016) 8:556-562.
pubpic2016chenWe report the development of a polymerase evolution system and its use to evolve thermostable polymerases that efficiently interconvert C2′-OMe modified oligonucleotides and their DNA counterparts via “transcription” and “reverse transcription,” or more importantly, PCR amplify partially C2′-OMe or C2′-F modified oligonucleotides.

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FRET characterization of complex conformational changes in a large 16S ribosomal RNA fragment site-specifically labeled using unnatural base pairs

T. Lavergne, R. Lamichhane, D.A. Malyshev, Z. Li, L. Li, E. Sperling, J.R. Williamson, D.P. Millar, F.E. Romesberg, ACS Chem. Biol. (2016) 11:1347-1353.
pubpic2016lavergneWe report the synthesis and evaluation of several unnatural ribotriphosphates bearing linkers that allow the chemoselective attachment of different functionalities. One unnatural base pair is used to dual label a 243-nt fragment of a 16S RNA with Cy3 and Cy5, which are then used to characterize conformational changes in the presence of ribosomal proteins.

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Temperature dependence of CN and SCN IR absorptions facilitates their interpretation and use as probes of proteins

R. Adhikary, J. Zimmermann, P.E. Dawson, F.E. Romesberg, Anal. Chem. (2015) 87:11561–11567.
pubpic2015badhikaryIn both model systems and the N-terminal Src homology 3 domain of Crk-II (nSH3), we show that the absorption frequency of cyano and thiocyano probes is linearly correlated with temperature and that the slope of the resulting line (frequency-temperature line slope or FTLS) reflects the nature of the probe’s microenvironment, including whether or not the probe is engaged in H-bonds.

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An alternative terminal step of the general secretory pathway in Staphylococcus aureus

A. Craney, M.M. Dix, R. Adhikary, B.F. Cravatt, F.E. Romesberg, mBio (2015) 6:e01178-15.
pubpic2015ccraneyWeidentify the S. aureus operon ayrRABC and show that once released from repression by AyrR, the protein products AyrABC together confer resistance to the SPase inhibitor arylomycin M131 by providing an alternate and novel method of releasing translocated proteins, and we demonstrate that AyrABC functionally complements SPase by mediating the processing of the normally secreted proteins. Overall, the data demonstrate that ayrRABC encodes a secretion stress-inducible alternate terminal step of the general secretory pathway.