SUPPLEMENTARY INFORMATION
The crystal structure of DNA mismatch repair protein MutS binding to a G:T
mismatch
Meindert H. Lamers*†, Anastassis Perrakis‡†, Jacqueline H. Enzlin*§, Herrie H. K.
Winterwerp*, Niels de Wind*||, Titia K. Sixma*
Figure A Surface charge representation48 of MutS dimer. Electrostatic charges (red
for negative, blue for positive) shown between –10 and +10 kT. The figure illustrates
the overall negative charge of MutS and the existence of channels through the protein.
The positive interface of the clamp domains with DNA is just visible, as well as an
additional positive groove at the top of the clamp domains.
Figure B Sequence alignment of E. coli MutS to human MSH2, MSH3 and MSH6.
The alignment used is the multi sequence alignment of ref 50. Secondary structure
elements (:-helix, : -strand, : 310-helix) are displayed and coloured according to
domain structure (see Fig. 2). Conservation displayed by grey boxes for similarity,
grey background for identity. Disordered loop indicated by blue dashed line under the
sequence. All DNA contacts (including van der Waals interactions) from the specific
mismatch-binding domain shown in blue background with white lettering. Nonspecific DNA binding residues denoted by light pink filled circles for mismatch
binding monomer, dark pink open circles for second monomer. All dimer contacts
from mismatch binding monomer in light green background, from second monomer in
dark green boxes. Sequence variants found in HNPCC patients (see Supplementary
information Fig. D) are indicated in red boxes in MSH2 and MSH6 sequences
respectively.
Figure C Domain movements between the two monomers of MutS, after
superposition of the core domains. A linear interpolation between the coordinates of
the two superimposed monomers was done in Cartesian coordinate space to create all
the (fictional) intermediate stages for the animation. The animation is intended only to
visualise differences in orientation of domains and does not claim to represent realtime movements. DNA and protein are represented as cartoon, coloured as in figure 2;
Phe36 and ADP as ball-and-stick models. Regions that are disordered in the nonmismatch binding monomer become transparent as the animation goes 'towards' that
monomer. The same is done for the ball-and-stick model of the ADP molecule which
is placed in a random place in the non-mismatch binding monomer to facilitate the
animation. The figure was drawn with BOBSCRIPT/Raster3D and animated with the
ImageMagick tools.
Figure D Missense mutations found in HNPCC families localised to MutS structure.
MSH6 mutations (green spheres) on mismatch-binding monomer (right), MSH2
mutations (blue spheres) on non mismatch binding monomer (left). Orientation as in
Fig. 1, colouring of backbone from grey to red with increasing conservation50.
Missense variants were assembled (June 2000) from the database of the international
collaborative group on HNPCC http://www.nfdht.nl/database/mdbchoice.htm and the
human gene mutation database http://www.uwcm.ac.uk/uwcm/mg/ns/1/203983.html.
MSH2 mutations: N127T, E198G, C199R, A305T, G322D, S323C, C333R L440P,
D506Y, R524P, E562V, P622L, A636P, H639R, E647K, G674D, G692R, C697F;
A834T. MSH6 mutations: S144I, S285I, Y556F, G566R, Q698E, D803G, Y850C,
P1087T/R, T1225M. See. Supplementary information Fig.B for mapping to MutS.
This mapping provides a basis for interpretation of additional functional studies that
will be required for definitive classification as polymorphism or pathogenic mutation.