RNA Aptamer selection against HA33 for Neurotoxin sequestion

Wesley Yu

RNA Aptamer Selection against HA33 for Neurotoxin sequestion

HA33 is a portion of three botulism neurotoxins secreted in a progenitor form from Clostridium botulinum. The HA33 protein can bind to glycoproteins that facilitate internal transportation through the bloodstream, and as such, can cause artery blockages leading to many circulatory problems associated with botulism, a unique but potentially fatal paralytic illness (Arndt et al. 2005). By binding to these glycolipids and glycoproteins, HA33 can cause flaccid muscle paralysis amongst other symptoms.

The HA33 protein occupies up to 30% of the progenitor complex, and acts as a dimer linking the entire progenitor complex with the other protein subunits HA17 and HA70. Although the HA33 protein complex’s specific structural role in the progenitor complex has not been properly defined yet, nucleic acids bind in general to the crystalline B-trefoil regions of HA33 (Arndt et al. 2005). Most of the other applications developed for this target revolve around its role in the progenitor structure, as well as its own complex crystalline structure. Some of these applications involve data collection about HA33’s relation to the other protein subunits in the progenitor complex, as well as HA33’s binding affiliation to carbohydrates.

Aptamers are oligonucleic acid or peptide molecules created from random sequence pools for the purpose of binding to specific targets. An aptamer developed for this protein would be useful in the medical field for its ability to prevent or relieve muscle paralysis by potentially changing the HA33 protein configuration. Consequently, new potential uses for the HA33 protein include applications in pharmaceutical drugs used for muscle paralysis and blood clotting. The aptamer would bind and induce a conformational change in HA33, preventing the binding of glycoproteins to the blood stream, allowing blood flow throughout the muscular and circulatory systems to be less restricted. Furthermore, the glycoproteins could be enchanced by the apatmer bounded HA33 to actually promote blood flow throughout the human circulatory system, if they bind to cells typically responsible for blood clots.

Specific Aim 1: Select an RNA aptamer against the HA33 protein that can be used to induce a conformational change in the target’s protein structure.

Figure 1: Progenitor toxin complex of C. botulinum comprised of the HA33 protein complex (Dark Blue), HA17 complex (Sky Blue) and HA70 complex (Yellow). The BoNT neurotoxin protein is indicated by the red segment, while the green segment indicates the NTNHA non-toxic, non-hemagglutinin protein. Adapted from Arndt (2005).

Target Order Information:

Vendor: Ellington Lab

Vendor Website: https://www.ellingtonlab.org

Central Lab Telephone: 512-471-6445

Office Manager: 512-232-3426

Cost per Unit: $0

Cost per Round: $0

References:

1.       Arndt JW, Gu J, Jaroszewski L, Schwarzenbacher R, Hanson MA, Lebeda FJ, Stevens RC. The Structure of the neurotoxin-associated protein HA33/A from Clostridium botulinum suggests a reoccurring beta-trefoil fold in the progenitor toxin complex, Molecular Biology 2005; 346 (4): 1083-1093.

2.       Nakamura T, Tonozuka T, Ito S, Takeda Y, Sato R, Matsuo I, Ito Y, Oguma K, Nishikawa A. Molecular diversity of the two sugar-binding sites of the Beta-trefoil lectin HA33/C (HA1) from Clostridium botulinum type C neurotoxin, Biochemistry and Biophysics 2011; 1,512 (1): 69-77.

3.       Van Damme EJ, Nakamura-Tsuruta S, Hirabayashi J, Rouge P, Peumans WJ. The Sclerotinia sclerotiorum agglutinin represents a novel family of fungal lectins remotely related to the Clostridium botulinum non-toxin haemagglutinin HA33/A, Molecular Biotechnology 2007; 24 (2-3): 143-156.

4.       Yamashita S, Yoshida H, Uchiyama N, Nakakita Y, Nakakita S, Tonozuka T, Oguma K, Nishikawa A, Kamitori S. Carbohydrate recognition mechanism of HA70 from Clostridium botulinum deduced from X-ray structures in complexes with sialylated oligosaccharides, Science Research and Medicine 2012; 586 (16): 2404-2410.

5.       Umeda K, Seto Y, Kohda T, Mukamoto M, Kozaki S. A novel multiplex PCR method for Clostridium botulinum neurotoxin type A gene cluster typing, Microbiology 2010; 54 (5): 308-312.

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