# Selection and sensing applications of DNAzymes selective for paramagnetic metal ions

> **NIH NIH R01** · UNIVERSITY OF ILLINOIS AT URBANA-CHAMPAIGN · 2020 · $257,422

## Abstract

Project summary / abstract
 The overall goal of this project is to develop and validate a novel class of fluorescent sensors for
paramagnetic metal ions (PMIs, e.g., Fe2+, Fe3+, Mn2+ and Mn3+), and to use these sensors to provide deeper
insight into the uptake and homeostasis of PMIs in bacteria and the role of PMIs in pathogenesis. PMIs are
essential elements for both humans and bacteria; the availability of these metal ions is sharply limited for
pathogens, as a part of a host defense mechanism known as “nutritional immunity”; the most well characterized
examples being Fe and Mn sequestration during infection. Moreover, Fe and Mn-regulated pathways are closely
linked with pathways involved in managing oxidative stress, as occurs in phagocytic respiratory burst. Despite
the importance of PMIs in nutritional immunity and oxidative stress pathways, the precise mechanisms dictating
nutritional immunity, bacterial uptake of PMIs, and the ability of certain bacterial strains to circumvent metal
starvation and thrive are unclear. A major barrier to understanding these complex mechanisms is the lack of
spatiotemporal detection of PMIs in their different OSs in living bacterial cells. This proposal seeks to overcome
this major barrier by selection and characterization of PMI-specific DNAzymes, and subsequent development
and validation of DNAzyme-based turn-on fluorescent sensors selective not only for different PMIs, but also
different oxidation states of the same PMI in two model systems (Staphylococcus aureus and Escherichia coli).
 Specifically, we plan to employ in vitro selection to obtain DNAzymes with high cleavage activity and strong
affinity for different PMIs (Fe2+ and Mn2+), while maintaining specificity for the different oxidation states of the
same metal ion (Fe2+ vs. Fe3+, and Mn2+ vs. Mn3+). Biochemical studies of these DNAzymes will provide
information about conserved sequences, pH and metal ion dependence, and kinetic parameters of the DNAzyme
activity. Biophysical characterization using spectroscopic methods (UV-vis and EPR) and x-ray crystallography
will elucidate PMI-binding stoichiometry, affinity and selectivity in these DNAzymes. The knowledge acquired will
be used to convert these DNAzymes into PMI sensors using the patented catalytic beacon technology. The use
of a “caged” and FRET DNAzyme sensor enabling quantitative monitoring of metal ion concentration and
speciation in living cells under temporal control will also be explored.
 Since pathogenic bacteria such as S. aureus and E. coli are a major public health issue, especially due to
the spread of antibiotic resistance, our ability to develop turn-on fluorescent sensors for the real time detection
of PMIs in cells will overcome a major barrier within the field of nutritional immunity by improving our
understanding of the uptake and homeostasis of PMIs in bacteria and the role of PMIs in pathogenesis.
Ultimately, knowledge gained from these sensors could provide insights neces...

## Key facts

- **NIH application ID:** 9908095
- **Project number:** 5R01GM124316-04
- **Recipient organization:** UNIVERSITY OF ILLINOIS AT URBANA-CHAMPAIGN
- **Principal Investigator:** Yi Lu
- **Activity code:** R01 (R01, R21, SBIR, etc.)
- **Funding institute:** NIH
- **Fiscal year:** 2020
- **Award amount:** $257,422
- **Award type:** 5
- **Project period:** 2017-07-15 → 2021-08-15

## Primary source

NIH RePORTER: https://reporter.nih.gov/project-details/9908095

## Citation

> US National Institutes of Health, RePORTER application 9908095, Selection and sensing applications of DNAzymes selective for paramagnetic metal ions (5R01GM124316-04). Retrieved via AI Analytics 2026-05-22 from https://api.ai-analytics.org/grant/nih/9908095. Licensed CC0.

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