CD133 aptamer

Jun 6, 2024 • Zhizhong Lu, Ying Ao

Description

In 2013, Shigdar, S. et al. performed iterative rounds of an in vitro selection process, known as the systematic evolution of ligands by exponential enrichment (SELEX), to identify RNA aptamers that specifically bind to CD133. Further studies identified one aptamer, CD133-A15, specifically bound to the same epitope as the AC133 antibody, while the other aptamer, CD133-B19, bound to the extracellular domain of the CD133 protein. These aptamers were efficiently internalised into CD133-positive cancer cells and showed superior penetration of three-dimensional tumour sphere^[1].

SELEX

Using Lipofectamine 2000, the C-terminally His-tagged CD133 was transfected into HEK293T cells and allowed to express for 72 h prior to SELEX experiments, with expression confirmed by AC133 antibody staining. Similar to previous SELEX experiments, a random RNA library of approximately 1 × 10¹⁴ species containing 2′-fluoro-modified ribose on all pyrimidines was incubated with HEK293T cells expressing human CD133. Unbound RNA was removed via several washing steps prior to RT-PCR, and the process was repeated for a total of 12 rounds^[1].
Detailed information are accessible on SELEX page.

Structure

The 2D structure of the figures are based on the article by ribodraw tool to draw^[1].

CD133-A15 aptamer: 5'-CCCUCCUACAUAGGG-3'
CD133-B19 aptamer: 5'-CAGAACGUAUACUAUUCUG-3'

Ligand information

SELEX ligand

CD133 antigen, also known as prominin-1, is a glycoprotein that in humans is encoded by the PROM1 gene. It is a member of pentaspan transmembrane glycoproteins, which specifically localize to cellular protrusions. When embedded in the cell membrane, the membrane topology of prominin-1 is such that the N-terminus extends into the extracellular space and the C-terminus resides in the intracellular compartment. The protein consists of five transmembrane segments, with the first and second segments and the third and fourth segments connected by intracellular loops while the second and third as well as fourth and fifth transmembrane segments are connected by extracellular loops.While the precise function of CD133 remains unknown, it has been proposed that it acts as an organizer of cell membrane topology.-----From Wiki

Name	Uniprot ID	Pfam	MW	Amino acids sequences	PDB	Gene ID
CD133	O43490	PF05478	97.202 KDa	MALVLGSLLLLGLCGNSFSGGQPSSTDAPKAWNYELPATNYETQDSHKAGPIGILFELVHIFLYVVQPRDFPEDTLRKFLQKAYESKIDYDKPETVILGLKIVYYEAGIILCCVLGLLFIILMPLVGYFFCMCRCCNKCGGEMHQRQKENGPFLRKCFAISLLVICIIISIGIFYGFVANHQVRTRIKRSRKLADSNFKDLRTLLNETPEQIKYILAQYNTTKDKAFTDLNSINSVLGGGILDRLRPNIIPVLDEIKSMATAIKETKEALENMNSTLKSLHQQSTQLSSSLTSVKTSLRSSLNDPLCLVHPSSETCNSIRLSLSQLNSNPELRQLPPVDAELDNVNNVLRTDLDGLVQQGYQSLNDIPDRVQRQTTTVVAGIKRVLNSIGSDIDNVTQRLPIQDILSAFSVYVNNTESYIHRNLPTLEEYDSYWWLGGLVICSLLTLIVIFYYLGLLCGVCGYDRHATPTTRGCVSNTGGVFLMVGVGLSFLFCWILMIIVVLTFVFGANVEKLICEPYTSKELFRVLDTPYLLNEDWEYYLSGKLFNKSKMKLTFEQVYSDCKKNRGTYGTLHLQNSFNISEHLNINEHTGSISSELESLKVNLNIFLLGAAGRKNLQDFAACGIDRMNYDSYLAQTGKSPAGVNLLSFAYDLEAKANSLPPGNLRNSLKRDAQTIKTIHQQRVLPIEQSLSTLYQSVKILQRTGNGLLERVTRILASLDFAQNFITNNTSSVIIEETKKYGRTIIGYFEHYLQWIEFSISEKVASCKPVATALDTAVDVFLCSYIIDPLNLFWFGIGKATVFLLPALIFAVKLAKYYRRMDSEDVYDDVETIPMKNMENGNNGYHKDHVYGIHNPVMTSPSQH	NA	8842

Some isolated sequences bind to the affinity of the protein.

Name	Sequence	Ligand	Affinity
CD133-A15 aptamer	5'-GGGCCAGGCAGCGAGGGUGUAUAGGUAUUGAAUCAAUUGUUUUAGCGUUGUCUUACCGACCACACGCGUCCGAGA-3'	CD133 (in Hep3B cell)	33.9 ± 12.5 nM
CD133-B19 aptamer	5'-GGGCCAGGCAGCGAGGGAACGCGAGCUUAGAUUUCGGAGAACAGUUUGUAAUUGUUCCGACCACACGCGUCCGAGA-3'	CD133 (in Hep3B cell)	52.3 ± 15.8 nM

References

[1] RNA aptamers targeting cancer stem cell marker CD133.
Shigdar, S., Qiao, L., Zhou, S. F., Xiang, D., Wang, T., Li, Y., Lim, L. Y., Kong, L., Li, L., & Duan, W.
Cancer letters, 330(1), 84–95. (2013)