Molecular requirements for degradation of a modified sense RNA strand by Escherichia coli ribonuclease H1
Bibliographic record
Abstract
The structural requirements for DNA/RNA hybrids to be suitable substrates for RNase H1 are well described; however the tolerance level of this enzyme towards modifications that do not alter the duplex conformation is not clearly understood, especially with respect to the sense RNA strand. In order to investigate the molecular requirements of Escherichia coli RNase H1 (termed RNase H1 here) with respect to the sense RNA strand, we synthesized a series of oligonucleotides containing 2'-deoxy-2'-fluoro-beta-D-ribose (2'F-RNA) as a substitute for the natural beta-D-ribose sugars found in RNA. Our results from a series of RNase H1 binding and cleavage studies indicated that 2'F-RNA/DNA hybrids are not substrates of RNase H1 and ultimately led to the conclusion that the 2'-hydroxyl moiety of the RNA strand in a DNA/RNA hybrid is required for both binding and hydrolysis by RNase H1. Through the synthesis of a series of chimeric sense oligonucleotides of mixed RNA and 2'F-RNA composition, the gap requirements of RNase H1 within the sense strand were examined. Results from these studies showed that RNase H1 requires at least five or six natural RNA residues within the sense RNA strand of a hybrid substrate for both binding and hydrolysis. The RNase H1-mediated degradation patterns of these hybrids agree with previous suggestions on the processivity of RNase H1, mainly that the binding site is located 5' to the catalytic site with respect to the sense strand. They also suggest, however, that the binding and catalytic domains of RNase H1 might be closer than has been previously suggested. In addition to the above, physicochemical studies have revealed the thermal stabilities and relative conformations of these modified heteroduplexes under physiological conditions. These findings offer further insights into the physical binding and catalytic properties of the RNase H1-substrate interaction, and have been incorporated into a general model summarizing the mechanism of action of this unique enzyme.
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How this classification was reachedexpand
Full frame machine prediction
Teacher imitationNot calibrated prevalence, not ground truth. Human validation pending. The Gemma side is a direct model label for every work in the frame, read from the title-only record. The Codex side is a classifier learned from the 10,348 direct Codex labels and calibrated to design-weighted sample rates; fields without enough sample support carry no Codex call. Candidate is the union of the two sides; consensus is their intersection. These outputs are machine_predicted_unvalidated and are not human labels.
Distilled classifier scores by category (both heads)
| Category | Codex | Gemma |
|---|---|---|
| Metaresearch | 0.000 | 0.001 |
| Meta-epidemiology (narrow) | 0.000 | 0.000 |
| Meta-epidemiology (broad) | 0.000 | 0.000 |
| Bibliometrics | 0.000 | 0.000 |
| Science and technology studies | 0.000 | 0.000 |
| Scholarly communication | 0.000 | 0.000 |
| Open science | 0.000 | 0.000 |
| Research integrity | 0.000 | 0.000 |
| Insufficient payload (model declined to judge) | 0.001 | 0.000 |
Machine scores (provisional)
The two teacher heads of the student model, read on this work. A score orders the frame for review; it never asserts a category, and the validation status ships verbatim with every row.
Baseline scores from an immature model (maturity gate not passed, 7 training rounds). Scores rank; they never assert a category.
score_only:v0-immature-baseline · verbatim from the scoring run: score_only means the number may rank works, and no category label ships from itClassification
machine, unvalidatedMachine predicted; a candidate call from one source (direct Gemma or distilled Codex), not a consensus.
How this classification was reached, model by model and score by score, is at the end of the page under "How this classification was reached".