Search

The Hammerhead Ribozyme (HHR) is a ubiquitous RNA enzyme that catalyzes site-specific intramolecular cleavage. While mutations to its catalytic core have traditionally been viewed as detrimental to its activity, several discoveries of naturally occurring variants of the full-length ribozyme challenge this notion, suggesting a deeper understanding of HHR evolution and functionality. By systematically introducing mutations at key nucleotide positions within the catalytic core, we generated single-, double-, and triple-mutation libraries to explore the sequence requirements and evolution of a full-length HHR. In vitro selection revealed many novel hammerhead variants, some of which possess mutations at nucleotides previously considered to be essential. We also demonstrate that the evolutionary trajectory of each nucleotide in the catalytic core directly correlates with their functional importance, potentially giving researchers a novel method to assess the sequence requirements of functional nucleic acids., (© 2024 The Authors. ChemBioChem published by Wiley-VCH GmbH.)

The 10-23 DNAzyme is one of the most active DNA-based enzymes, and in theory, can be designed to target any purine-pyrimidine junction within an RNA sequence for cleavage. However, purine-pyrimidine junctions within a large, structured RNA (lsRNA) molecule of biological origin are not always accessible to 10-23, negating its general utility as an RNA-cutting molecular scissor. Herein, we report a generalizable strategy that allows 10-23 to access any purine-pyrimidine junction within an lsRNA. Using three large SARS-CoV-2 mRNA sequences of 566, 584 and 831 nucleotides in length as model systems, we show that the use of antisense DNA oligonucleotides (ASOs) that target the upstream and downstream regions flanking the cleavage site can restore the activity (kobs) of previously poorly active 10-23 DNAzyme systems by up to 2000-fold. We corroborated these findings mechanistically using in-line probing to demonstrate that ASOs reduced 10-23 DNAzyme target site structure within the lsRNA substrates. This approach represents a simple, efficient, cost-effective, and generalizable way to improve the accessibility of 10-23 to a chosen target site within an lsRNA molecule, especially where direct access to the genomic RNA target is necessary., (© The Author(s) 2024. Published by Oxford University Press on behalf of Nucleic Acids Research.)

Background: The minority of working-age Finns eat according to the national and Nordic nutritional guidelines and increasing numbers of health problems affect the Finnish workforce. Coincidently recruiting new workers in the more rural areas of Finland, such as Satakunta, has been problematic. To optimize the use of the existing workforce, health promotion interventions focusing on nutrition have been suggested to improve the health and well-being of the current working age Finns., Methods and Analysis: The aim of this RCT study is to assess the effectiveness of a 12-month multifactorial nutritional guidance intervention to improve work ability (performance), work well-being, health-related quality of life, work productivity, sickness absence, dietary intake and eating habits. In total, six small or medium-sized companies and their employees (n = 170) from the Satakunta region will be recruited. Companies will be randomized 1:1 to a 12-month multifactorial nutritional guidance intervention group (INT) or a control group (CG). Comprehensive measurements are taken before randomization (baseline) and at the end of the 12-month study period. Primary outcomes (work ability, work well-being and health-related quality of life) are measured with Work Ability Index, Utrecht Work Engagement Scale short questionnaire and EQ-5D. Dietary intake and eating habits are measured with 3-day food records and Food Frequency Questionnaire (FFQ)., Discussion: This study will provide nationally important data on how workplace nutrition guidance affects work-related outcomes, quality of life, and nutritional and overall health status among working age Finns., Competing Interests: Declaration of competing interest The authors declare no conflict of interest. The funders have no role in the design of the study., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Systematic evolution of ligands by exponential enrichment (SELEX) has been used to discover thousands of aptamers since its development in 1990. Aptamers are short single-stranded oligonucleotides capable of binding to targets with high specificity and selectivity through structural recognition. While aptamers offer advantages over other molecular recognition elements such as their ease of production, smaller size, extended shelf-life, and lower immunogenicity, they have yet to show significant success in real-world applications. By analyzing the importance of structured library designs, reviewing different SELEX methodologies, and the effects of chemical modifications, we provide a comprehensive overview on the production of aptamers for applications in drug delivery systems, therapeutics, diagnostics, and molecular imaging., (© 2024 The Authors. Angewandte Chemie International Edition published by Wiley-VCH GmbH.)

A new method for the detection of genomic RNA combines RNA cleavage by the 10-23 DNAzyme and use of the cleavage fragments as primers to initiate rolling circle amplification (RCA). 230 different 10-23 DNAzyme variants were screened to identify those that target accessible RNA sites within the highly structured RNA transcripts of SARS-CoV-2. A total of 28 DNAzymes were identified with >20 % cleavage, 5 with >40 % cleavage and one with >60 % in 10 min. The cleavage fragments from these reactions were then screened for coupling to an RCA reaction, leading to the identification of several cleavage fragments that could efficiently initiate RCA. Using a newly developed quasi-exponential RCA method with a detection limit of 500 aM of RNA, 14 RT-PCR positive and 15 RT-PCR negative patient saliva samples were evaluated for SARS-CoV-2 genomic RNA, achieving a clinical sensitivity of 86 % and specificity of 100 % for detection of the virus in <2.5 h., (© 2023 The Authors. Chemistry - A European Journal published by Wiley-VCH GmbH.)