Ivy Mead
24 March 2011
Bioinformatics
Lab report 4
Three-finger toxins (3FTx) have been found in several different groups of vertebrates,
including snakes (elapidae) as well as mammals (Naimuddin et al., 2011). In some venomous
snakes 3FTx proteins have been found in the venom as widely effective neurotoxins transmitted
by bite. As well as wide ranging neurological effects, 3FTx proteins have also been known to
have cytotoxic and platelet inhibition effects (Jiang et al., 2011). 3FTx proteins in general have
been found to imitate interleukins and targets for ion channels and receptors in the body, which
is made possible by it’s molecular structure. The 3FTx proteins are characterized by a highly
conserved cysteine structure and a highly diverse series of loop regions. The variation in the loop
regions is thought to aid in molecular mimicry (Naimuddin et al., 2011).
Thus eleven sequences of distinct 3FTx proteins isolated from snake venom were aligned
and a set of analyses was carried out in order to answer several interesting questions. An
evolutionary tree of the sequences was produced, providing a general structure to the data
(Figure 1). Interestingly enough, one species (Ophiophagus Hannah) had two different types of
3FTx proteins and they were placed on different branches of the tree. Literature sources state that
evolution of the toxin family 3FTx is driven by positive selection, which makes sense to a certain
extent (Jiang et al., 2011). Toxic venom can aid a snake in two ways, by allowing them to
disable prey more easily and by aiding in protection against potential predators. The more
effective the venom is at doing those things, likely the better the snake is expected to adapt.
However, the positive selection pressures only seem to be affecting the second region of the
protein, as seen by looking at the mRNA sequence. When aligned, the first half of the sequence
is highly conserved and the second half is very diverse. The sequence diversity caused by
positive selection in that second half is likely the key to the success of the protein because it
allows a wide range of harmful effects to the prey and/or predator of the snake. However, as with
everything in evolutionary biology, there could always be other factors playing a part that could
be obscuring the true causes in this case.
To explore the possibility of forces of selection shaping the evolution of the 3FTx toxic
protein family, codon-based tests of neutral, positive, and purifying selection were carried out in
MEGA using the Nei-Gojobori method on the data collected. The results of the codon-based
tests of neutrality carried out showed a very small number of statistically significant probabilities
(P < 0.05) indicating neutral selection acting on the protein family, thus it is unclear if that force
is acting on the family. The results of the codon-based tests of positive selection showed also a
small number of statistically significant probabilities (P < 0.05) indicating that neutral selection
could possibly be acting on the protein family but not for certain. The results of the codon-based
tests of purifying selection showed no statistically significant probabilities (P < 0.05) indicating
that it is unlikely that purifying selection has been a driving force for the evolution of the protein
family (Tamura et al, 2007) (Nei and Gojobori, 1986).
Part of what seems to make the 3FTx toxins so effective as a part of venom is their ability
to bind to many different receptors and neurons by being able to mimic the binding site. The first
half of nucleotide sequence coding for the protein was found to be highly conserved, likely a
remnant of the family’s pre-venom structure, which allows it to bind to many different sites in
the prey’s body. The part that seems to distinguish the protein from related proteins, is the
second half of the sequence, which as mentioned earlier is highly variable and most likely
ultimately allows the protein to be toxic.
Figure 1. An evolutionary relationship tree of the studied 3FTx sequences from different
types of snakes. Units of tree are in number of base substitutions per site. This figure was
compiled using UPGMA method, with distances calculated using the Juke-Cantor method
using the program MEGA5 (Sneath and Sokal, 1973)(Jukes and Cantor, 1969).
Works Cited:
Jiang, Y. et al., Venom gland transcriptomes of two elapid snake (Bungarus multicinctus and
Naja atra) and evolution of toxin genes. 2011. BMC Genomics. 12: 1. Accessed on Mar
17, 2011 from http://www.biomedcentral.com/1471-2164/12/1.
Jukes, T. and C. Cantor. Evolution of protein molecules. In Munto, H., editor. Mammalian
Protein Metabolism. Academic Press: New York. 1969.
Nei, M. and T. Gojobori. Simple methods for estimating the numbers of synonymous and
nonsynonymous nucleotide substitutions. 1986. Molecular Biology and Evolution, 3:418426.
Naimuddin, M., et al. Directed evolution of a three-finger neurotoxin by using cDNA display
yields antagonists as well as agonists of interleukin-6 receptor signaling. 2011. Molecular
Brain. 4: 2. Accessed Mar 17, 2011 from http://www.molecularbrain.com/content/4/1/2.
Sneath, P. and R. Sokal. Numerical Taxonomy. Freeman: San Francisco. 1973.
Tamura K. et al., MEGA4: Molecular Evolutionary Genetics Analysis (MEGA) software version
4.0. 2007. Molecular Biology and Evolution 24:1596-1599.