Title

Site-directed mutagenesis studies of M1-toxin1: Amino acids responsible for toxin-M1 receptor interaction

Date of Award

2000

Availability

Article

Degree Name

Doctor of Philosophy (Ph.D.)

First Committee Member

Lincoln Potter, Committee Chair

Abstract

The venom of the green mamba Dendroaspis angusticeps contains several m1-isotoxins with similar amino acid sequences and muscarinic subtype pharmacology. The most prevalent isotoxin, m1-toxin1, has total selectivity for M1 receptors over M2-M5, binds irreversibly to M1 receptors and can bind to M1 receptors occupied with 3H-NMS.There is striking homology between the m1-isotoxins and the remaining 11 muscarinic toxins that bind reversibly to muscarinic receptors. This thesis studied some of the amino acids in m1-toxin1 which are different from those in the reversible toxins. These residues were mutated from the amino acid present in the m1-toxins to the analogous residue in the common muscarinic toxins, and each mutant was examined for its selectivity for M1 receptors, reversibility of binding and allosteric effects.The cDNA for m1-toxin1 was cloned from the venom glands of Dendroaspis angusticeps. The full length cDNA was 514 bp long and included the ubiquitous signal peptide present in all cloned short-chain neurotoxins. The cDNA was expressed in Pichia pastoris and the recombinant toxin behaved as identical to native toxin in binding assays. to M1-M5 receptors.

Keywords

Biology, Molecular; Health Sciences, Toxicology; Health Sciences, Pharmacology

Link to Full Text

http://access.library.miami.edu/login?url=http://gateway.proquest.com/openurl?url_ver=Z39.88-2004&rft_val_fmt=info:ofi/fmt:kev:mtx:dissertation&res_dat=xri:pqdiss&rft_dat=xri:pqdiss:9972561