Home
Database
Research
About us
Lys1.2minF aptamer, Lys1.2minE apatmer
Description
In 2013, Camille S Padlan and Vladimir N Malashkevich used the SELEX method to isolate the aptamer with high compatibility for the HEWL(lysozyme). At the same time, they examined its structure by X-ray diffrcation. It has a sequence length of 59. Camille S Padlan and Vladimir N Malashkevich systematically minimized an RNA aptamer (Lys1) selected against hen egg white lysozyme. The resultant 59-nucleotide compact aptamer (Lys1. 2minE) retains nanomolar binding affinity and the ability to inhibit lysozyme’s catalytic activity. Lys1. 2minE inhibits catalysis of large cell wall substrates but not catalysis of small model substrates. Then, Camille S Padlan and Vladimir N Malashkevich shortened the terminal stem of Lys1. 2minE to four Watson-Crick base pairs and got the Lys1. 2minF. At the same time, they examined its structure by X-ray diffrcation. It has a sequence length of 45. Its structure may be an adaptable protein binding platform. It can bind lysozyme with high affinity. It inhibits lysozyme cleavage of large substrates, and it bounds aptamer has no effect on catalysis of small substrates[1].SELEX
RNA aptamers were internally labeled with [α32P]-GTP and allowed to bind to lysozyme for 1 h at room temperature. The bound complex was passed through a dual filter dot blot system attached to a vacuum manifold. The nitrocellulose membrane captures the aptamer–protein complexes while unbound aptamers are trapped by the nylon filter directly beneath. Both filters are visualized by phosphor storage imaging, and the fraction protein bound was calculated by determining the volume of radioactivity retained on the nitrocellulose divided by the total radioactivity retained by both filters. Sedimentation velocity experiments were performed on a Beckman XL-I using a standard procedure. Determining the S20, and D20, values of the RNA aptamers under each buffer condition specified by analytical ultracentrifugation. A standard lysozyme activity assay was used to determine the enzymatic activity[1].
Detailed information are accessible on SELEX page.
Structure
2D representation
Here we use ribodraw to complete the figure, through the 3D structure information[1].
Lys1.2minF: 5'-GGGCGGCUAAAGAGUGCAGAGUUACUUAGUUCACUGCAGACGCCC-3'
Lys1.2minE: 5'-GGGUUCAUCAGGGCUAAAGAGUGCAGAGUUACUUAGUUCACUGCAGACUUGACGAACCC-3'
|
|
3D visualisation
Camille S Padlan and Vladimir N Malashkevich sovled the crystal structure, at 2.68 A resolution, of an RNA aptamer bound to Lysozyme C protein has been determined. The PDB ID of this structure is 4M6D[1].Additional available structures that have been solved and detailed information are accessible on Structures page.
(Clicking the "Settings/Controls info" to turn Spin off)
|
|
Camille S Padlan and Vladimir N Malashkevich sovled the crystal structure, at 2.68 A resolution, of an RNA aptamer bound to Lysozyme C protein has been determined. The PDB ID of this structure is 4M4O[1].
Additional available structures that have been solved and detailed information are accessible on Structures page.
(Clicking the "Settings/Controls info" to turn Spin off)
|
|
Binding pocket
Left: Surface representation of the binding pocket of the aptamer generated from PDB ID: 4M6D at 2.68 Å resolution. Lysozymes(shown in vacuumm electrostatics), blue is positive charge, red is negative charge. Right: The hydrogen bonds of binding sites of the aptamer bound with Lysozymes.
|
|
Left: Surface representation of the binding pocket of the aptamer generated from PDB ID: 4M4O at 2 Å resolution. Lysozymes(shown in vacuumm electrostatics), blue is positive charge, red is negative charge. Right: The hydrogen bonds of binding sites of the aptamer bound with Lysozymes.
|
|
Ligand information
SELEX ligand
Camille S Padlan and Vladimir N Malashkevich distinguished structures Lys1. 2 and Lys1. 3 by trimming nucleotides predicted by each to be largely unstructured. Removing different nucleotides to test whether it can binding with lysozyme, and testing its affinity[1].| Name | Sequence | Ligand | Affinity |
|---|---|---|---|
| Lys1. 2minF | 5'-GGGCGGCUAAAGAGUGCAGAGUUACUUAGUUCACUGCAGACGCCC-3' | HEWL(Hen Egg White Lysozyme) | 57±3nM |
| Lys1. 2minE | 5'-GGGUUCAUCAGGGCUAAAGAGUGCAGAGUUACUUAGUUCACUGCAGACUUGACGAACCC-3' | HEWL(Hen Egg White Lysozyme) | 19±2nM |
Structure ligand
Lysozymes have primarily a bacteriolytic function; those in tissues and body fluids are associated with the monocyte-macrophage system and enhance the activity of immunoagents. Has bacteriolytic activity against M. luteus. Lysozyme C is capable of both hydrolysis and transglycosylation; it shows also a slight esterase activity. It acts rapidly on both peptide-substituted and unsubstituted peptidoglycan, and slowly on chitin oligosaccharides.-----from uniprot
| Uniprot ID | Pfam | MW | Amino acids sequences | PDB ID | GenBank |
|---|---|---|---|---|---|
| P00698 | PTHR11407 | 14.31 kDa | KVFGRCELAAAMKRHGLDNYRGYSLGNWVCAAKFESNFNTQATNRNTDGSTDYGILQINSRWWCNDGRTPGSRNLCNIPCSALLSSDITASVNCAKKIVSDGNGMNAWVAWRNRCKGTDVQAWIRGCRL | 4wm6 | 9031 |
 |
Similar compound
We used the Dail server website to compare the structural similarities of ligand proteins, and chose the top 10 in terms of similarity for presentation. The Dali server is a network service for comparing protein structures in 3D. Dali compares them against those in the Protein Data Bank (PDB). Z-score is a standard score that is converted from an original score. The list of neighbours is sorted by Z-score. Similarities with a Z-score lower than 2 are spurious. RMSD(Root Mean Square Deviation) value is used to measure the degree to which atoms deviate from the alignment position.
| PDB | Z-score | RMSD | Description |
|---|---|---|---|
| 1LSG-A | 23 | 1 | Hen Egg White Lysozyme |
| 1NF5-C | 20.6 | 1.4 | Alpha-Lactalbumin |
| 4OWD-A | 8.1 | 2.7 | Membrane-Bound Lytic Murein Transglycosylase f |
| 4C5F-A | 8 | 2.5 | Membrane-Bound Lytic Murein Transglycosylase c |
| 6TA9-A | 7.9 | 2.5 | Slt Domain-Containing Protein |
| 1QTE-A | 7.9 | 2.6 | Soluble Lytic Transglycosylase Slt70 |
| 3BKH-A | 7.8 | 2.8 | Lytic Transglycosylase |
| 5E27-A | 7.8 | 2.5 | Resuscitation-Promoting Factor Rpfb |
| 4YIM-B | 7.7 | 2.8 | Putative Soluble Lytic Murein Transglycosylase |
| 7LAM-A | 7.5 | 3.1 | Lytic Transglycosylase Domain-Containing Protein |
References
[1] An RNA aptamer possessing a novel monovalent cation-mediated fold inhibits lysozyme catalysis by inhibiting the binding of long natural substrates.Padlan, C. S., Malashkevich, V. N., Almo, S. C., Levy, M., Brenowitz, M., & Girvin, M. E.
RNA, 20(4), 447–461. (2013)