RNA aptamer as potential antidote against botulism: an in vivo report

Janardhanan, Pavithra; Ravichandran, Easwaran; Cai, Shuowei

doi:10.1504/tbj.2015.078135

Cited by 2 publications

(2 citation statements)

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“…The endopeptidase activity of fLCA and tLCA with endogenous SNAP-25 in the cytosol of neuroblastoma cells was studied by delivering the LCAs inside the cytosol. Cytosolic delivery of 50 nM fLCA or tLCA was achieved using Lipofectamine 2000 (Life Technologies, Grand Island, NY); control cells received no LCA treatment 59 , 60 .…”

Section: Methodsmentioning

confidence: 99%

A novel role of C-terminus in introducing a functionally flexible structure critical for the biological activity of botulinum neurotoxin

Feltrup

Patel

Kumar³

et al. 2018

Sci Rep

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Botulinum neurotoxin (BoNT) is responsible for botulism, a clinical condition resulting in flaccid muscle paralysis and potentially death. The light chain is responsible for its intracellular toxicity through its endopeptidase activity. Available crystal structures of BoNT/A light chains (LCA) are based on various truncated versions (tLCA) of the full-length LCA (fLCA) and do not necessarily reflect the true structure of LCA in solution. The understanding of the mechanism of action, longevity of intoxication, and an improved development of endopeptidase inhibitors are dependent on first having a better insight into the structure of LCA in solution. Using an array of biophysical techniques, we report that the fLCA structure is significantly more flexible than tLCA in solution, which may be responsible for its dramatically higher enzymatic activity. This seems to be achieved by a much stronger, more rapid binding to substrate (SNAP-25) of the fLCA compared to tLCA. These results suggest that the C-terminus of LCA plays a critical role in introducing a flexible structure, which is essential for its biological function. This is the first report of such a massive structural role of the C-terminus of a protein being critical for maintaining a functional state.

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Section: Methodsmentioning

confidence: 99%

A novel role of C-terminus in introducing a functionally flexible structure critical for the biological activity of botulinum neurotoxin

Feltrup

Patel

Kumar³

et al. 2018

Sci Rep

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“…In our earlier studies, by using radioactive based SELEX approach, we have successfully identified three 2’-fluoro-pyrimidines modified RNA aptamers against light chain of botulinum neurotoxin type A (LCA) (27), which not only showed high binding affinity and high specificity to LCA, but also showed strong inhibition effects on enzymatic activity of LCA, and in vivo protection of BoNT intoxication (28). Surface plasmon resonance (SPR) has been developed as an alternative for selection of aptamers (29, 30).…”

Section: Introductionmentioning

confidence: 99%

Selection of RNA Aptamers Against Botulinum Neurotoxin Type A Light Chain Through a Non-Radioactive Approach

Chang

Janardhanan

Mello

et al. 2016

Appl Biochem Biotechnol

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Botulinum neurotoxin (BoNT), a category A agent, is the most toxic molecule known to mankind. The endopeptidase activity of light chain domain of BoNT is the cause for the inhibition of the neurotransmitter release and the flaccid paralysis that leads to lethality in botulism. Currently, antidotes are not available to reverse the flaccid paralysis caused by BoNT. In the present study, a non-radioactive based SELEX process is developed by utilizing surface plasmon resonance to monitor the binding enrichment. Two RNA aptamers have been identified as strong binders against light chain of botulinum neurotoxin type A. These two aptamers showed strong inhibition activity on LCA, with IC50 in nM range. Inhibition kinetic studies reveal mid nanomolar KI and non-competitive nature of their inhibition, suggesting they have strong potential as antidotes that can reverse the symptom caused by BoNT/A. More importantly, we observed that 2′-fluorine-pyrimidines modified RNA aptamers identified here do not change their binding and biological activities. This observation could lead to a cost-effective way for Systematic Evolution of Ligands by EXponential enrichment (SELEX), by using regular nucleotide during SELEX, and 2′-fluorine-pyrimidines modified nucleotide for final application to enhance their RNase-resistance.

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RNA aptamer as potential antidote against botulism: an in vivo report

Cited by 2 publications

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A novel role of C-terminus in introducing a functionally flexible structure critical for the biological activity of botulinum neurotoxin

A novel role of C-terminus in introducing a functionally flexible structure critical for the biological activity of botulinum neurotoxin

Selection of RNA Aptamers Against Botulinum Neurotoxin Type A Light Chain Through a Non-Radioactive Approach

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