Supplementary Figure Legends - Word file (35 KB )

Manuscript # 2004-07-21763A
Supplementary Figure Legends
Figure captions
Figure S1
Structural validation of the trapping strategy and crosslinking
biochemistry. a, Close-up view of the intrahelical region surrounding the site of
crosslink introduction. The extrahelical active site lies outside this view, to the
middle right of each panel. Top left, the original lesion-recognition complex1
used in design of the trapping strategy, with the hydrogen bond highlighted
between Asn149 and N4 of the estranged C (refer also to Fig. 1c, left); top right,
the trapped complex of hOGG1-QC (this work) with oxoG-containing DNA; and
bottom, the trapped complex of N149C-hOGG1 with undamaged DNA (this
work). Shown in dark blue in the crosslinked structures is a A-weighted2 Fo-Fc
map contoured at 3, leaving out the atoms of the thiol-bearing tether attached
to the estranged C (-CH2-CH2S-). The weak density for the hydrocarbon portion
of the crosslink is indicative of high local conformational mobility. b, Nonreducing SDS-PAGE gel monitoring progress of disulfide crosslink formation
between hOGG1-QC (see text) and DNA presenting oxoG or G to the active
site, as a function of time. c, Selectivity in formation of crosslinked complex
between N149C hOGG1 and G-containing DNA, as a function of tether length
(See Fig. 1c for a structural representation of n).
Fig. S2. A weighted2 2Fo-Fc electron density maps contoured at 1 shown
around selected atoms of the G-complex in the regions of (a) the extrahelical G
and (b) estranged C. The DNA backbone is in gold, G in red, estranged C in
magenta, and key residues of the protein in cyan. Density for the His270 side
chain at the lower left hand corner of (a) was weak.
Fig. S3. Structural representation of the expected interactions between the
main-chain carbonyl Gly42 and nucleobases examined in this study. Whereas
oxoG is known to hydrogen bond with Gly42, the lone pair of electrons on G
engender a repulsive interaction with the lone pairs on Gly42. Neither repulsive
nor attractive interactions occur with 7-deazaG or 7-deaza-8-aza-G.
Fig. S4. A weighted2 2Fo-Fc maps contoured at 1 around selected atoms of
the lesion-recognition pocket in the structures of the (a) 7-deaza-8-azaGcomplex and (b) 7-deazaG-complex (region shown and view similar to righthand panels in Fig. 5).
Bruner, S. D., Norman, D. P. & Verdine, G. L. Structural basis for recognition
and repair of the endogenous mutagen 8-oxoguanine in DNA. Nature 403, 85966 (2000).
Read, R. J. Improved Fourier coefficients for maps using phases from partial
structures with errors. Acta Crystallogr. A 42, 104-149 (1986).