Molecular Genetics

Soft shell clams (Mya arenaria) are bivalves native to the east coast of North America. Some individual clams are resistant to a harmful neurotoxin called saxitoxin while others are not. Resistance results from a mutation in the gene encoding a voltage gated sodium channel, a transmembrane protein in the axon of neuron cells. Neurons are affected by saxitoxin because the toxin blocks voltage gated sodium channels along nerve axons preventing nerve signals from being transmitted from the dendrite to the axon terminus.

amino acid readout of the affected gene with the single mutation highlighed

DNA, Central Dogma and the Voltage Gated Sodium Channel

DNA is made up of long strings of nucleotides. There are four nucleotides in DNA: Adenine (A), Thymine (T), Guanine (G) and Cytosine (C). These nucleotides are held together by a phosphate-sugar “backbone” and are complemented by a parallel strand of nucleotides making a DNA molecule “double-stranded”. Parallel strands are held together by hydrogen bonds between bases; Adenine binds to Thymine, Cytosine binds to Guanine. Discrete sections of DNA carry sequences of nucleotides that, when processed, are responsible for proteins or functional RNA chains. These sections are called genes. When making a protein, the DNA is first transcribed into a single-stranded piece of messenger RNA (mRNA) using an enzyme called RNA polymerase. The newly made mRNA is then translated through a ribosome into a long chain of amino acids. This chain of amino acids folds up to make a functional protein. Proteins build the organism, including all measurable and observable characteristics.

Diagram of Glutamic acid, coded for by GAA or GAG, and Aspartic acid, coded for by GAU or GAC.

Substitution Mutation

There are two alleles for the gene that codes for the voltage gated sodium channel protein in soft shell clams: E and D. In clams sensitive to saxitoxin paralysis, the amino acid at position 632 in the sodium channel protein is a glutamic acid (E). However, in clams resistant to saxitoxin paralysis, the amino acid at position 632 in the sodium channel protein is aspartic acid (D) [note: this numbering is based on partial gene sequence of the sodium channel]. The nucleotide codon that results in a glutamic acid can be either GAA or GAG whereas the nucleotide codon that results in aspartic acid can be either GAT or GAC. This means that saxitoxin resistance in soft shell clams can be attributed to one single nucleotide substitution mutation. Functionally, aspartic acid is shorter than glutamic acid by one methylene group. The presence or absence of this one methylene group is the difference between being able to bind saxitoxin or not, and hence whether or not the clam is sensitive or resistant to saxitoxin.

Mutation and the Activation Gate

The voltage gated sodium channel protein has four transmembrane domains with six subdomains in each. The space between subdomains 5 and 6 form a pore that allows sodium passage through the membrane into the axon. The extracellular loops between subdomains 5 and 6 form the activation gate. When the polarity of the cell becomes negative, the voltage gated sodium channel changes shape (conformation) conformation causing the extracellular gate to open. Some of the amino acid residues in the extra-cellular loops between these subdomains form active sites that allow for saxitoxin binding. The mutation and subsequent change from glutamic acid to aspartic acid at amino acid position #632 in the sodium channel protein changes the physical properties of one of the binding sites located in domain II, between sub-domains 5 and 6. This changes the binding affinity for the entire pore making the clam resistant to saxitoxin.