@article{stuart_koshlap_guenther_agris_2003, title={Naturally-occurring modification restricts the anticodon domain conformational space of tRNA(Phe)}, volume={334}, ISSN={["1089-8638"]}, DOI={10.1016/j.jmb.2003.09.058}, abstractNote={Post-transcriptional modifications contribute chemistry and structure to RNAs. Modifications of tRNA at nucleoside 37, 3'-adjacent to the anticodon, are particularly interesting because they facilitate codon recognition and negate translational frame-shifting. To assess if the functional contribution of a position 37-modified nucleoside defines a specific structure or restricts conformational flexibility, structures of the yeast tRNA(Phe) anticodon stem and loop (ASL(Phe)) with naturally occurring modified nucleosides differing only at position 37, ASL(Phe)-(Cm(32),Gm(34),m(5)C(40)), and ASL(Phe)-(Cm(32),Gm(34),m(1)G(37),m(5)C(40)), were determined by NMR spectroscopy and restrained molecular dynamics. The ASL structures had similarly resolved stems (RMSD approximately 0.6A) of five canonical base-pairs in standard A-form RNA. The "NOE walk" was evident on the 5' and 3' sides of the stems of both RNAs, and extended to the adjacent loop nucleosides. The NOESY cross-peaks involving U(33) H2' and characteristic of tRNA's anticodon domain U-turn were present but weak, whereas those involving the U(33) H1' proton were absent from the spectra of both ASLs. However, ASL(Phe)-(Cm(32),Gm(34),m(1)G(37),m(5)C(40)) exhibited the downfield shifted 31P resonance of U(33)pGm(34) indicative of U-turns; ASL(Phe)-(Cm(32),Gm(34),m(5)C(40)) did not. An unusual "backwards" NOE between Gm(34) and A(35) (Gm(34)/H8 to A(35)/H1') was observed in both molecules. The RNAs exhibited a protonated A(+)(38) resulting in the final structures having C(32).A(+)(38) intra-loop base-pairs, with that of ASL(Phe)-(Cm(32),Gm(34),m(1)G(37),m(5)C(40)) being especially well defined. A single family of low-energy structures of ASL(Phe)-(Cm(32),Gm(34), m(1)G(37),m(5)C(40)) (loop RMSD 0.98A) exhibited a significantly restricted conformational space for the anticodon loop in comparison to that of ASL(Phe)-(Cm(32),Gm(34),m(5)C(40)) (loop RMSD 2.58A). In addition, the ASL(Phe)-(Cm(32),Gm(34),m(1)G(37),m(5)C(40)) average structure had a greater degree of similarity to that of the yeast tRNA(Phe) crystal structure. A comparison of the resulting structures indicates that modification of position 37 affects the accuracy of decoding and the maintenance of the mRNA reading frame by restricting anticodon loop conformational space.}, number={5}, journal={JOURNAL OF MOLECULAR BIOLOGY}, author={Stuart, JW and Koshlap, KM and Guenther, R and Agris, PF}, year={2003}, month={Dec}, pages={901–918} }
 @article{sengupta_vainauskas_yarian_sochacka_malkiewicz_guenther_koshlap_agris_2000, title={Modified constructs of the tRNA T Psi C domain to probe substrate conformational requirements of m(1)A(58) and m(5)U(54) tRNA methyltransferases}, volume={28}, ISSN={["0305-1048"]}, DOI={10.1093/nar/28.6.1374}, abstractNote={The TPsiC stem and loop (TSL) of tRNA contains highly conserved nucleoside modifications, m(5)C(49), T(54), Psi(55)and m(1)A(58). U(54)is methylated to m(5)U (T) by m(5)U(54)methyltransferase (RUMT); A(58)is methylated to m(1)A by m(1)A(58)tRNA methyltransferase (RAMT). RUMT recognizes and methylates a minimal TSL heptadecamer and RAMT has previously been reported to recognize and methylate the 3'-half of the tRNA molecule. We report that RAMT can recognize and methylate a TSL heptadecamer. To better understand the sensitivity of RAMT and RUMT to TSL conformation, we have designed and synthesized variously modified TSL constructs with altered local conformations and stabilities. TSLs were synthesized with natural modifications (T(54)and Psi(55)), naturally occurring modifications at unnatural positions (m(5)C(60)), altered sugar puckers (dU(54)and/or dU(55)) or with disrupted U-turn interactions (m(1)Psi(55)or m(1)m(3)Psi(55)). The unmodified heptadecamer TSL was a substrate of both RAMT and RUMT. The presence of T(54)increased thermal stability of the TSL and dramatically reduced RAMT activity toward the substrate. Local conformation around U(54)was found to be an important determinant for the activities of both RAMT and RUMT.}, number={6}, journal={NUCLEIC ACIDS RESEARCH}, author={Sengupta, R and Vainauskas, S and Yarian, C and Sochacka, E and Malkiewicz, A and Guenther, RH and Koshlap, KM and Agris, PF}, year={2000}, month={Mar}, pages={1374–1380} }
 @article{koshlap_guenther_sochacka_malkiewicz_agris_1999, title={A distinctive RNA fold: The solution structure of an analogue of the yeast tRNA(Phe) T psi C domain}, volume={38}, ISSN={["0006-2960"]}, DOI={10.1021/bi990118w}, abstractNote={The structure of an analogue of the yeast tRNAPhe T Psi C stem-loop has been determined by NMR spectroscopy and restrained molecular dynamics. The molecule contained the highly conserved modification ribothymidine at its naturally occurring position. The ribothymidine-modified T Psi C stem-loop is the product of the m5U54-tRNA methyltransferase, but is not a substrate for the m1A58-tRNA methyltransferase. Site-specific substitutions and 15N labels were used to confirm the assignment of NOESY cross-peaks critical in defining the global fold of the molecule. The structure is unusual in that the loop folds far over into the major groove of the curved stem. This conformation is stabilized by both stacking interactions and hydrogen bond formation. Furthermore, this conformation appears to be unique among RNA hairpins of similar size. There is, however, a considerable resemblance to the analogous domain in the crystal structure of the full-length yeast tRNAPhe. We believe, therefore, that the structure we have determined may represent an intermediate in the folding pathway during the maturation of tRNA.}, number={27}, journal={BIOCHEMISTRY}, author={Koshlap, KM and Guenther, R and Sochacka, E and Malkiewicz, A and Agris, PF}, year={1999}, month={Jul}, pages={8647–8656} }