Ribostamycin aptamer, Paromomycin aptamer

Mar 17, 2025 • Jiali Wang, Ke Chen

Timeline

Description

In 2008, Suess, B. et al. developed a two-stage strategy of in vitro selection followed by a genetic screen and identified several artificial small molecule-binding riboswitches that respond to the aminoglycoside neomycin. They display no sequence similarities to in vitro selected neomycin aptamers but contain parts of the decoding site that is the binding site for neomycin on the ribosomal RNA. In 2016, Wöhnert, J. et al. explored the mechanism of ligand discrimination (neomycin, paromomycin, ribostamycin) by neomycin synthetic riboswitches at atomic resolution and determined the structures of the 27 nucleotides engineered neomycin sensing riboswitch RNA-ribostamycin complex and the neomycin sensing riboswitch RNA bound to paromomycin using NMR, but failed to resolve the structure of the neomycin synthetic riboswitch^[1,5].

SELEX

In 2008, Suess, B. et al. used an in vitro selection (Systematic Evolution of Ligands by EXponential enrichment, SELEX) method and in vivo genetic screen to identify neomycin riboswitches controlling translation initiation. They enriched an RNA pool for neomycin B binding through six in vitro selection cycles. This pool had a 74-nucleotide random region between constant parts and was inserted before a gfp reporter gene in a yeast vector, generating 5 × 10⁴ sequences for in vivo screening. Yeast cells were transformed and screened in two steps: first for gene expression without ligand, then for neomycin-dependent regulation. This yielded 30 candidates with neomycin-dependent fluorescence decrease. Sequence analysis found 10 unique candidates with a fully conserved 16-nucleotide sequence in the top two and partially in others. The sequences of Ribomycin aptamer and Paromycin aptamer come from here^[1].

Detailed information are accessible on SELEX page.

Structure

2D representation

In 2008, Suess, B. et al. used SELEX and in vivo genetic screen to obtain 10 unique candidates with a fully conserved 16-nucleotide sequence in the top two and partially in others. The sequences of these two aptamers were obtained by truncating N1 from 10 unique candidates. Here we used ribodraw to complete the figure, through the 3D structure information. The aptamer was initially isolated for neomycin and was named by Suess, B^[1].

5'-GGCUGCUUGUCCUUUAAUGGUCCAGUC-3'

3D visualisation

The solution structure of the 27 nucleotides engineered neomycin sensing riboswitch RNA-ribostamycin complex was determined by Suess, B. et al. through multidimensional NMR spectroscopy. The PDB ID of this structure is 2N0J^[5].

Additional available structures that have been solved and detailed information are accessible on Structures page.

The solution structure of the neomycin sensing riboswitch RNA bound to paromomycin was determined by Michael Famulok et al. through multidimensional NMR spectroscopy. The PDB ID of this structure is 2MXS^[5].

Additional available structures that have been solved and detailed information are accessible on Structures page.

Binding pocket

Left: Surface representation of the binding pocket of the aptamer generated from PDB ID: 2N0J by NMR. Ribostamycin (shown in sticks) is labeled in magenta. Right: The hydrogen bonds of binding sites of the aptamer bound with ribostamycin.

Left: Surface representation of the binding pocket of the aptamer generated from PDB ID: 2MXS by NMR. Paromomycin (shown in sticks) is labeled in magenta. Right: The hydrogen bonds of binding sites of the aptamer bound with paromomycin.

Ligand information

SELEX ligand

Suess, B. & Wöhnert, J. used isothermal titration calorimetry (ITC) to measure the affinity between the artificial riboswitches (aptamer) and paromycin, ribomycin and neomycin^[5].

Structure ligand

An antibiotic used to treat a wide variety of bacterial infections in the body.-----From Drugbank

PubChem CID: a unique identifier for substances in the PubChem database.

CAS number: a global registry number for chemical substances.

Drugbank: a comprehensive database with detailed information on drugs and drug targets.

Name	PubChem CID	Molecular Formula	Molecular Weight	CAS	Solubility	Drugbank ID
Ribostamycin	33042	C17H34N4O10	454.5 g/mol	25546-65-0	10 mM	DB03615

An antibiotic used to treat complications of liver failure and infections caused by parasites in the intestines.-----From Drugbank

Name	PubChem CID	Molecular Formula	Molecular Weight	CAS	Solubility	Drugbank ID
Paromomycin	165580	C23H45N5O14	615.6 g/mol	7542-37-2	7.97e + 01 g/L	DB01421

Similar compound(s)

We screened the compounds with great similarity to ribostamycin and paromomycin by using the ZINC database and showed some of the compounds' structure diagrams. For some CAS numbers not available, we will supplement them with Pubchem CID.

ZINC ID: a compound identifier used by the ZINC database, one of the largest repositories for virtual screening of drug-like molecules.

PubChem CID: a unique identifier for substances in the PubChem database.

CAS number: a global registry number for chemical substances.

ZINC ID	Name	CAS	Pubchem CID	Structure
ZINC8214383	Dibekacin	34493-98-6	470999
ZINC4095654	Neamine	3947-65-7	72392
ZINC253673980	Lividomycin	36441-41-5	72394
ZINC8214590	Kanamycin	59-01-8	6032
ZINC8214692	Tobramycin	32986-56-4	36294
ZINC85536952	Framycetin	119-04-0	8378
ZINC53132258	Kanamycin B	4696-76-8	439318

References

[1] Screening for engineered neomycin riboswitches that control translation initiation.
Weigand, J. E., Sanchez, M., Gunnesch, E. B., Zeiher, S., Schroeder, R., & Suess, B.
RNA (New York, N.Y.), 14(1), 89–97. (2008)
[2] NMR resonance assignments of an engineered neomycin-sensing riboswitch RNA bound to ribostamycin and tobramycin.
Schmidtke, S. R., Duchardt-Ferner, E., Weigand, J. E., Suess, B., & Wöhnert, J.
Biomolecular NMR assignments, 4(1), 115–118. (2010)
[3] Highly modular structure and ligand binding by conformational capture in a minimalistic riboswitch.
Duchardt-Ferner, E., Weigand, J. E., Ohlenschläger, O., Schmidtke, S. R., Suess, B., & Wöhnert, J.
Angewandte Chemie (International ed. in English), 49(35), 6216–6219. (2010)
[4] Sequence-specific inhibition of Dicer measured with a force-based microarray for RNA ligands.
Limmer, K., Aschenbrenner, D., & Gaub, H. E.
Nucleic acids research, 41(6), e69. (2013)
[5] What a Difference an OH Makes: Conformational Dynamics as the Basis for the Ligand Specificity of the Neomycin-Sensing Riboswitch.
Duchardt-Ferner, E., Gottstein-Schmidtke, S. R., Weigand, J. E., Ohlenschläger, O., Wurm, J. P., Hammann, C., Suess, B., & Wöhnert, J.
Angewandte Chemie (International ed. in English), 55(4), 1527–1530. (2016)
[6] Synthetic Dual-Input Hybrid Riboswitches─Optimized Genetic Regulators in Yeast.
Kelvin, D., Arias Rodriguez, J., Groher, A. C., Petras, K., & Suess, B.
ACS synthetic biology, 14(2), 497–509. (2025)