C. elegans Tc3 Transposase Protein
Niki Watson '01 and Ashley Rowatt '03
Transposition is a method of recombination in which
an element of DNA is excised and inserted into a new location in the genome.
The element of DNA that moves is called the transposon, and there are a
number of classes defining different types of elements. The most
understood eukaryotic transposon families are Tc1/mariner and Ac/hobo.
In true Molecular Biology humor, the transposons are often given creative
names such as pogo and Tigger, both of which belong to the
Tc1/mariner family (Smitt and Riggs, 1996).
In addition to containing normal genetic information,
members of the Tc1/mariner family code for a single protein, transposase,
which catalyzes the transposition. In order to catalyze the movement,
tranposase must recognize, excise, and relocate the transposon (Puoderoyen
al., 1997). The mechanism of recognition involves several specific
DNA-protein interactions which are the focus of this tutorial.
II. General Structure
Transposase exists in dimer
form <DNA binding domain
of the protein is of particular importnace <
III. DNA Binding Domain
elegans Tc3 transposase binding domain has been crystallized with DNA
(van Pouderoyen et al., 1997). One monomer
binds one molecule of DNA <first
helix of each monomer is involved in the dimerazation
of the protein <helix-turn-helix
motif is a highly conserved amino acid structure which is commonly found
in DNA-protein interactions (Wintjens and Rooman, 1996). A well known
example of a protein that exhibits the HTH motif is the lambda repressor
(Weaver, 1999). This motif allows the protein to make substantial
interactions with the DNA. The N-terminus
of the transposase protein is involved in specific DNA binding <
IV. Major Groove Interactions
The HTH motif makes extensive
contacts with the major groove of the DNA. Helix 2 and helix 3 each
make specific bonds to the DNA molecule, thus allowing the transposase
to recognize specific DNA sequences. The DNA-protein interactions
consist of hydrogen bonding and salt bridges.
Helix 2 interacts extensively
with the phosphate backbone of the major groove. However, the helix
makes one specific contact with a purine: His26
hydrogen bonds to G7
hydrogen bonds to phosphate group 6
with phosphate groups 5
and 6 <
Helix 3 also bonds with
elements of the major groove. Specifically, Arg36
hydrogen bonds to G8
interacts with T9
through a thymine-specific hydrogen bond via a molecule of water
also form salt bridges with the phosphate group 8
IV. Minor Groove Interactions
In addition to major groove
interactions, base-specific recognition of the transposon is furthered
by interactions within the minor groove. The majority of the minor
groove contacts result from binding with the transposase N-terminus
and Arg3 insert
themselves into the minor groove <
Smit, A.F., A.B. Riggs. 1996. Tiggers and other
DNa transposon fossils in the human genome. Proc. Natl. Acad.
Sci. USA. 93, 1443-1448.
G., R. Ketting, A. Perrakis, R. Plasterk, and T. Sixma. 1997.
Crystal structure of the specific DNA-binding
domain of Tc3 transposase of C. elegans in complex with transposon
DNA. EMBO J.
Weaver, R.F. 1999. Molecular Biology. McGraw-Hill,
Boston, 788 pp.
R., and M. Rooman. 1996. Structural classification of HTH
DNA-binding domains and protein-DNA interaction modes. J. Mol. Biol.262,