Nucleic Acid Aptamer Selection against HA-33 of C. botulinum for an Early Diagnosis and a Treatment for Botulism

Nucleic Acid Aptamer Selection against HA-33 of C. botulinum for an Early Diagnosis and a Treatment for Botulism

Sharon Joy

       Botulism, an uncommon yet fatal illness characteristically manifests as an acute paralysis that begins with the cranial nerves and moves toward the lower extremities is caused by the rod-shaped and spore-forming bacterium Clostridium botulinum. The neurotoxins produced by this certain bacterium are among some of the most toxic bacterial toxins ever known. This protein neurotoxin interrupts or inhibits the neuromuscular transmission that leads to paralysis. The bacterium produces seven neurotoxins in which four of them affect humans. There are different kinds of botulism which include: food-borne botulism, wound botulism, infant botulism, adult infectious botulism and inadvertent botulism (World Health Organization). Treatment of this illness includes botulinus antitoxins which inhibits the function of the toxins in the blood stream. However, early diagnosis and treatment of this illness is key, since delayed treatment can lead to an advance in paralysis in patients including respiratory muscle paralysis which can lead to a patient requiring ventilatory care, and ICU care; not diagnosing early enough could also lead to death in patients (MedlinePlus Medical Encyclopedia).

       A botulinum toxin complex consists of a neurotoxic, a nontoxic nonhaemagglutinin and a haemagglutinin or the HA complex. The HA complex consists of arms which include two HA-33 arms. A study was conducted on the absorption of the botulinum toxin in rat intestinal epithelial cell line, which showed that cell binding and monolayer transport of serotype D toxins significantly increased with an increase of HA-33/HA-17 arms. This led to the conclusion that the HA-33/HA-17 arm was not required yet it was a vital component in facilitating the transport of the toxins (Federation of European Microbiological Societies). HA-33 is also crucial for toxin movement, activation and protection (Journal of Molecular Biology). A selected aptamer against HA33 can be used for an early diagnosis of this fatal illness and can be modified for a therapeutic application. This aptamer can be tagged with a molecule that can result in a change of shape for HA33, which results in an inhibition of the function of the protein. By changing the shape of the protein, it can obstruct the protein from entering through the microvilli to the small intestine membrane. By inhibiting the protein’s function, which is responsible for botulism, it can in turn help in preventing the fatal disease that leads to paralysis.

Figure 1: This figure depicts the large botulinum complex which consists of 

a neurotoxic (BoNT), a nontoxic nonhaemagglutinin (NTHA) and a haemagglutinin (HA) complex with three components: HA-33, HA-17 and HA-70. The HA-33/HA-17 arms greatly facilitate the transportation of the toxins. By selecting an aptamer against HA-33, which has an unique organization and a carbohydrate recognition site, it can be modified for a therapeutic application which can inhibit the toxin from entering the intestinal membrane (InTech).

Specific Aim 1: The identification and amplification of a RNA aptamer selected against HA-33. 

Specific Aim 2: Modification of the aptamer for a therapeutic application. By tagging the aptamer with a molecule that changes the shape of the protein target when it binds to it, it can inhibit the function of the paralysis responsible for the fatal illness. 

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

Arndt, J. W., J. Gu, R. Schwarzenbacher, M. A. Hanson, F. J. Lebeda, and R. C. Stevens. "The Structure of the Neurotoxin-associated Protein HA33/A from Clostridium Botulinum Suggests a Reoccurring β-Trefoil Fold in the Progenitor Toxin Complex." Journal of Molecular Biology 4th ser. 346.4 (2005): 1083-093. 

"Botulism: Treatment." MedlinePlus Medical Encyclopedia. U.S. National Library of Medicine, 2012. Web. 03 Sept. 2013.

"Clostridium Botulinum." International Programme on Chemical Safety Poisons Information Monograph858 Bacteria. World Health Organization, n.d. Web. 02 Sept. 2013. <http://www.who.int/csr/delibepidemics/clostridiumbotulism.pdf>.

Ito, H., Y. Sagane, K. Miyata, K. Inui, T. Matsuo, R. Horiuchi, and T. Ikeda. "HA-33 Facilitates Transport of the Serotype D Botulinum Toxin across a Rat Intestinal Epithelial Cell Monolayer." Federation of European Microbiological Societies (2011): n. pag. 

Sagane, Y., K. Inui, S. Miyashita, K. Miyata, K. Niwa, T. Watanabe, and T. Suzuk. "Botulinum Toxin Complex: A Delivery Vehicle of Botulinum Neurotoxin Traveling Digestive Tract." InTech. N.p., 22 Aug. 2012. Web.

Proposal Link: https://docs.google.com/file/d/0B8u9l0gKo2jddHZkbHFSdWFmTW8/edit?usp=sharing
Link to first report: https://docs.google.com/file/d/0B8u9l0gKo2jdcmZFUUJ5NjlNTVk/edit?usp=sharing