C. elegans Tc3 Transposase Protein

Niki Watson '01 and Ashley Rowatt '03

Chime Index

Contents:

I. Introduction

    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 et 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  <>, in which each monomer binds a separate transposon end.  Since the recognition of transposon DNA is essential to the process of transposition, the DNA binding domain of the protein is of particular importnace <>.


III.  DNA Binding Domain

    The  C. elegans Tc3 transposase binding domain has been crystallized with DNA (van Pouderoyen et al., 1997).    One monomer binds one molecule of DNA <>.  The monomer consists of three alpha helices.  The first helix of each monomer is involved in the dimerazation of the protein <> .  The second and third helices of each monomer form a helix-turn-helix (HTH) motif <>and participate in binding the transposon DNA.  The HTH 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 <>(see below).



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 <>.  The remainder of the helix contacts are with the phosphate backbone.  For example, Ser24 hydrogen bonds to phosphate group 6 <>.  Additionally, salt bridges link Arg30 with phosphate groups 5 and 6  <>.
    Helix 3 also bonds with elements of the major groove.  Specifically, Arg36 hydrogen bonds to G8 <>. Arg40 interacts with T9 through a thymine-specific hydrogen bond via a  molecule of water <>.  Both Arg36 and Arg40 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 <>.  The side chains of Pro2 and Arg3 insert themselves into the minor groove  <> and make extensive interactions with the DNA.  Additionally, it is believed that the transposase C-terminus connects with the minor groove, however, these interactions can not be detected in the crystal structure.


V. References
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.

van Pouderoyen, 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. 16, 6044-6054.

Weaver, R.F.  1999.  Molecular Biology.  McGraw-Hill, Boston, 788 pp.

Wintjens, R., and M. Rooman.  1996.  Structural classification of HTH DNA-binding domains and protein-DNA interaction modes. J. Mol. Biol.262, 294-313.