Solution structure and functional importance of a conserved RNA hairpin of eel LINE UnaL2

The eel long interspersed element (LINE) UnaL2 and its partner short interspersed element (SINE) share a conserved 3′ tail that is critical for their retrotransposition. The predicted secondary structure of the conserved 3′ tail of UnaL2 RNA contains a stem region with a putative internal loop. Deletion of the putative internal loop region abolishes UnaL2 mobilization, indicating that this putative internal loop is required for UnaL2 retrotransposition; the exact role of the putative internal loop in retrotransposition, however, has not been elucidated. To establish a structure-based foundation on which to address the issue of the putative internal loop function in retrotransposition, we used NMR to determine the solution structure of a 36 nt RNA derived from the 3′ conserved tail of UnaL2. The region forms a compact structure containing a single bulged cytidine and a U–U mismatch. The bulge and mismatch region have conformational flexibility and molecular dynamics simulation indicate that the entire stem of the 3′ conserved tail RNA can anisotropically fluctuate at the bulge and mismatch region. Our structural and mutational analyses suggest that stem flexibility contributes to UnaL2 function and that the bulged cytidine and the U–U mismatch are required for efficient retrotransposition

Real-world management of treatment-naïve diabetic macular oedema : 2-year visual outcome focusing on the starting year of intervention from STREAT-DMO study

Background/aims To investigate the yearly change of real-world outcomes for best corrected visual acuity (BCVA) after 2-year clinical intervention for treatment-naïve diabetic macular oedema (DMO). Methods Retrospective analysis of aggregated, longitudinal medical records obtained from 27 retina specialised institutions in Japan from Survey of Treatment for DMO database. A total of 2049 treatment-naïve centre involving DMO eyes of which the initial intervention started between 2010 and 2015, and had been followed for 2 years, were eligible. As interventions, antivascular endothelial growth factor (VEGF) agents, local corticosteroids, macular photocoagulation and vitrectomy were defined. In each eye, baseline and final BCVA, the number of each intervention for 2 years was extracted. Each eye was classified by starting year of interventional treatment. Results Although baseline BCVA did not change by year, 2-year improvement of BCVA had been increased, and reached to +6.5 letters in the latest term. There is little difference among starting year about proportions of eyes which BCVA gained >15 letters, in contrast to those which lost >15 letters were decreased by year. The proportion of eyes receiving anti-VEGF therapy was dramatically increased, while those receiving the other therapies were gradually decreased. The proportion of eyes which maintained socially good vision of BCVA>20/40 has been increased and reached to 59.0% in the latest term. Conclusion For recent years, treatment patterns for DMO have been gradually but certainly changed; as a result, better visual gain, suppression of worsened eyes and better final BCVA have been obtained. Anti-VEGF therapy has become the first-line therapy and its injection frequency has been increasing

Real-world management of treatment-naïve diabetic macular oedema in Japan : two-year visual outcomes with and without anti-VEGF therapy in the STREAT-DME study

Background/Aims To investigate real-world outcomes for best-corrected visual acuity (BCVA) after 2-year clinical intervention for treatment-naïve, centr-involving diabetic macular oedema (DME). Methods Retrospective analysis of longitudinal medical records obtained from 27 institutions specialising in retinal diseases in Japan. A total of 2049 eyes with treatment-naïve DME commencing intervention between 2010 and 2015 who were followed for 2 years were eligible. Interventions for DME included anti-vascular endothelial growth factor (VEGF) therapy, local corticosteroid therapy, macular photocoagulation and vitrectomy. Baseline and final BCVA (logMAR) were assessed. Eyes were classified by the treatment pattern, depending on whether anti-VEGF therapy was used, into an anti-VEGF monotherapy group (group A), a combination therapy group (group B) and a group without anti-VEGF therapy (group C). Results The mean 2-year improvement of BCVA was −0.04±0.40 and final BCVA of >20/40 was obtained in 46.3% of eyes. Based on the treatment pattern, there were 427 eyes (20.9%) in group A, 807 eyes (39.4%) in group B and 815 eyes (39.8%) in group C. Mean improvement of BCVA was −0.09±0.39, –0.02±0.40 and −0.05±0.39, and the percentage of eyes with final BCVA of >20/40 was 49.4%, 38.9%, and 52.0%, respectively. Conclusion Following 2-year real-world management of treatment-naïve DME in Japan, BCVA improved by 2 letters. Eyes treated by anti-VEGF monotherapy showed a better visual prognosis than eyes receiving combination therapy. Despite treatment for DME being selected by specialists in consideration of medical and social factors, a satisfactory visual prognosis was not obtained, but final BCVA remained >20/40 in half of all eyes

Conformational plasticity of RNA for target recognition as revealed by the 2.15 Å crystal structure of a human IgG–aptamer complex

Aptamers are short single-stranded nucleic acids with high affinity to target molecules and are applicable to therapeutics and diagnostics. Regardless of an increasing number of reported aptamers, the structural basis of the interaction of RNA aptamer with proteins is poorly understood. Here, we determined the 2.15 Å crystal structure of the Fc fragment of human IgG1 (hFc1) complexed with an anti-Fc RNA aptamer. The aptamer adopts a characteristic structure fit to hFc1 that is stabilized by a calcium ion, and the binding activity of the aptamer can be controlled many times by calcium chelation and addition. Importantly, the aptamer–hFc1 interaction involves mainly van der Waals contacts and hydrogen bonds rather than electrostatic forces, in contrast to other known aptamer–protein complexes. Moreover, the aptamer–hFc1 interaction involves human IgG-specific amino acids, rendering the aptamer specific to human IgGs, and not crossreactive to other species IgGs. Hence, the aptamer is a potent alternative for protein A affinity purification of Fc-fusion proteins and therapeutic antibodies. These results demonstrate, from a structural viewpoint, that conformational plasticity and selectivity of an RNA aptamer is achieved by multiple interactions other than electrostatic forces, which is applicable to many protein targets of low or no affinity to nucleic acids

Thermodynamic study of aptamers binding to their target proteins

International audienc

Development of the 12-Base Short Dimeric Myogenetic Oligodeoxynucleotide That Induces Myogenic Differentiation

A myogenetic oligodeoxynucleotide (myoDN), iSN04 (5′-AGA TTA GGG TGA GGG TGA-3′), is a single-stranded 18-base telomeric DNA that serves as an anti-nucleolin aptamer and induces myogenic differentiation, which is expected to be a nucleic acid drug for the prevention of disease-associated muscle wasting. To improve the drug efficacy and synthesis cost of myoDN, shortening the sequence while maintaining its structure-based function is a major challenge. Here, we report the novel 12-base non-telomeric myoDN, iMyo01 (5′-TTG GGT GGG GAA-3′), which has comparable myogenic activity to iSN04. iMyo01 as well as iSN04 promoted myotube formation of primary-cultured human myoblasts with upregulation of myogenic gene expression. Both iMyo01 and iSN04 interacted with nucleolin, but iMyo01 did not bind to berberine, the isoquinoline alkaloid that stabilizes iSN04. Nuclear magnetic resonance revealed that iMyo01 forms a G-quadruplex structure despite its short sequence. Native polyacrylamide gel electrophoresis and a computational molecular dynamics simulation indicated that iMyo01 forms a homodimer to generate a G-quadruplex. These results provide new insights into the aptamer truncation technology that preserves aptamer conformation and bioactivity for the development of efficient nucleic acid drugs.ArticleBioTech 2024, 13(2), 11journal articl

NMR structures of double loops of an RNA aptamer against mammalian initiation factor

doi:10.1093/nar/gki22

Structural and molecular basis for hyperspecificity of RNA aptamer to human immunoglobulin G

Potential applications for functional RNAs are rapidly expanding, not only to address functions based on primary nucleotide sequences, but also by RNA aptamer, which can suppress the activity of any target molecule. Aptamers are short DNA or RNA folded molecules that can be selected in vitro on the basis of their high affinity for a target molecule. Here, we demonstrate the ability of RNA aptamers to recognize—and bind to—human IgG with high specificity and affinity. An optimized 23-nucleotide aptamer, Apt8-2, was prepared, and was shown to bind to the Fc domain of human IgG, but not to other IgG's, with high affinity. Apt8-2 was observed to compete with protein A, but not with the Fcγ receptor, for IgG binding. NMR chemical-shift analyses localized the aptamer-binding sites on the Fc subdomain, which partially overlaps the protein A binding site but not the Fcγ receptor binding site. The tertiary structures of the predicted recognition sites on the Fc domain differ significantly between human IgG and other species of IgGs; this, in part, accounts for the high specificity of the selected aptamer. Apt8-2 can therefore be used as a protein A alternative for affinity purification of human IgG and therapeutic antibodies. Using Apt8-2 would have several potential advantages, raising the possibility of developing new applications based on aptamer design