# Single Molecule Studies of Nucleic Acids Remodeling

> **NIH NIH R35** · JOHNS HOPKINS UNIVERSITY · 2020 · $360,250

## Abstract

Project Summary
Double stranded DNA formed via Watson-Crick basepairing needs to be separated into single strands for
genome duplication or repair to occur. Likewise, double stranded RNA are frequently separated into single
strands during transcription, splicing, ribosome biogenesis and translation. Such nucleic acids unwinding
should not occur indiscriminately. Using advanced single molecule measurement technologies we discovered
a novel mechanism of regulating helicase activities through a conformational switch. This led to the
development of a superhelicase that can unwind thousands of base pairs processively even against a strong
opposing force.
A mirror process to nucleic acids unwinding is protein-dependent annealing of two stands of nucleic acids.
Examples include Rec/Rad51-mediated DNA recombination, non-coding RNA-based gene regulation and
CRISPR-based DNA degradation. All of these processes rely on basepairing interactions above the threshold
number of bp for specificity. How the target DNA or RNA can be both rapidly and accurately identified in the
presence of other sequences in large excess is an unanswered question.
`Nucleic acids remodeling' mediated by proteins either in the direction of unwinding or in the direction of
annealing is the overarching theme of this project. The premise here is that a balance between the basepair
breaking and formation is critical in normal functions of these proteins inside the cell and if the balance is not
properly maintained, it leads to mis-regulation and diseased states. The key questions to address are: (1) What
is the in vivo role of various helicase conformations? (2) Can we detect all reaction intermediates during
helicase function? (3) Can we mimic co-transcriptional RNA folding and ribosome assembly using
superhelicases? (4) How do basepairing interactions control replication fork reversal and restoration by
annealing helicases? (5) How do basepairing interactions determine in vivo kinetics of sRNA-based gene
regulation? (6) How does the balance between heteroduplex extension and reversal control CRISPR-
Cas9/Cpf1 target verification and cleavage activation? (7) How does the balance between heteroduplex
extension and reversal control Cas3 helicase-nuclease recruitment?

## Key facts

- **NIH application ID:** 9924561
- **Project number:** 5R35GM122569-04
- **Recipient organization:** JOHNS HOPKINS UNIVERSITY
- **Principal Investigator:** Taekjip Ha
- **Activity code:** R35 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2020
- **Award amount:** $360,250
- **Award type:** 5
- **Project period:** 2017-05-01 → 2022-04-30

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/9924561

## Citation

> US National Institutes of Health, RePORTER application 9924561, Single Molecule Studies of Nucleic Acids Remodeling (5R35GM122569-04). Retrieved via AI Analytics 2026-05-26 from https://api.ai-analytics.org/grant/nih/9924561. Licensed CC0.

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