Most RNA folding studies have been performed under non-physiological conditions of high concentrations (≥10 mM) of Mg2+free, while actual cellular concentrations of Mg2+free are only ~1 mM in a background of greater than 50 mM Mg2+total. To uncover cellular behavior of RNA, we devised cytoplasm mimic systems that include biological concentrations of amino acids, which weakly chelate Mg2+. Amino acid-chelated Mg2+ (aaCM) of ~15 mM dramatically increases RNA folding and prevents RNA degradation. Furthermore, aaCM enhance self-cleavage of several different ribozymes, up to 100,000-fold at Mg2+free of just 0.5 mM, indirectly through RNA compaction. Other metabolites that weakly chelate magnesium offer similar beneficial effects, which implies chelated magnesium may enhance RNA function in the cell in the same way. Overall, these results indicate that the states of Mg2+ should not be limited to free and bound only, as weakly bound Mg2+ strongly promotes RNA function under cellular conditions.

Background: Topologically knotted tRNA methyltransferases specifically recognize substrate tRNA. Results: Site-directed mutagenesis studies, chimeric protein analysis, and pre-steady state kinetics clarify the tRNA recognition sites of TrmH. Conclusion:The N-and C-terminal regions function in the initial binding process, and substrate tRNA is discriminated by the catalytic domain in an induced-fit process. Significance: Study of how proteins recognize RNA is crucial for understanding RNA maturation processes.

Sodiation−desodiation behaviors were investigated for electrodes composed of various binary phosphides, InP, CuP 2 , GeP, SiP, and LaP, as anode materials of a Na-ion battery. Although LaP electrode did not react with Na, the other electrodes showed reversible sodiation− desodiation reactions in the initial cycles. Rapid capacity decays were observed for CuP 2 , GeP, and SiP electrodes. In contrast, a better cyclability was attained for the InP electrode. These results indicate that binary phosphides (M−P) require four properties for improving cyclability: (i) low thermodynamic stability of M−P, (ii) high electronic conductivity of M, (iii) low hardness of M, and (iv) reactivity of M with Na.

RNA structure plays roles in myriad cellular events including transcription, translation, and RNA processing. Genome-wide analyses of RNA secondary structure in vivo by chemical probing have revealed critical structural features of mRNAs and long ncRNAs. Here, we examine the in vivo secondary structure of a small RNA class, tRNAs. Study of tRNA structure is challenging because tRNAs are heavily modified and strongly structured. We introduce “tRNA structure-seq,” a new workflow that accurately determines in vivo secondary structures of tRNA. The workflow combines dimethyl sulfate (DMS) probing, ultra-processive RT, and mutational profiling (MaP), which provides mutations opposite DMS and natural modifications thereby allowing multiple modifications to be identified in a single read. We applied tRNA structure-seq to E. coli under control and stress conditions. A leading folding algorithm predicts E. coli tRNA structures with only ∼80% average accuracy from sequence alone. Strikingly, tRNA structure-seq, by providing experimental restraints, improves structure prediction under in vivo conditions to ∼95% accuracy, with more than 14 tRNAs predicted completely correctly. tRNA structure-seq also quantifies the relative levels of tRNAs and their natural modifications at single nucleotide resolution, as validated by LC-MS/MS. Our application of tRNA structure-seq yields insights into tRNA structure in living cells, revealing that it is not immutable but has dynamics, with partial unfolding of secondary and tertiary tRNA structure under heat stress that is correlated with a loss of tRNA abundance. This method is applicable to other small RNAs, including those with natural modifications and highly structured regions.

To date, numerous modified nucleosides in tRNA as well as tRNA modification enzymes have been identified not only in thermophiles but also in mesophiles. Because most modified nucleosides in tRNA from thermophiles are common to those in tRNA from mesophiles, they are considered to work essentially in steps of protein synthesis at high temperatures. At high temperatures, the structure of unmodified tRNA will be disrupted. Therefore, thermophiles must possess strategies to stabilize tRNA structures. To this end, several thermophile-specific modified nucleosides in tRNA have been identified. Other factors such as RNA-binding proteins and polyamines contribute to the stability of tRNA at high temperatures. Thermus thermophilus, which is an extreme-thermophilic eubacterium, can adapt its protein synthesis system in response to temperature changes via the network of modified nucleosides in tRNA and tRNA modification enzymes. Notably, tRNA modification enzymes from thermophiles are very stable. Therefore, they have been utilized for biochemical and structural studies. In the future, thermostable tRNA modification enzymes may be useful as biotechnology tools and may be utilized for medical science.