List of Tables and Figures
Table 1. Parameters used to adjust the roll angles to bend the DNA: .
Base pair numbering: I1 starts at n = 1; I2 starts at n =
1; spacer starts at n = 18. Sequence “spacer-ext” consists of the 4-bp
spacer region between I1 and I2 and the first two bp ofI2 .
Figure 1. 560-bp amplicon comprised of the araI1 and araI2half-sites. The black region contains the sequence for I1 –
spacer – I2 : tagcatttttatccata agattagcggatcctacctga in
which the half-sites are underlined. I1 is on the left. The white
region precedes I1-spacer-I2 and has a length of 392 bp. The
striped region has a length of 130 bp. The 302-bp amplicon is similar
except that I1-spacer-I2 is located closer to the left end. Refer
to Figure S1 for the full sequence information.
Figure 2. Surface representations of DBD-DNA docking. N-terminus of DBD
(white) and 5’ end of DNA (gray) are shown by the black regions. (A)
Initial docking of DBD/I1 . A large gap exists. (B) DBD/I1complex after bending the DNA to reduce the gap. (C) DBD/I2 after
slight bending of DNA.
Figure 3. AFM images of samples prepared with DNA only and samples with
DNA, AraC, and arabinose. More sharply bent DNA is visible in samples
prepared with AraC and arabinose. The circular features contain protein.
Width of image = 500 nm. (A) 302 bp DNA only, (B and C) 302 bp
DNA+AraC+arabinose. (D) 560 bp DNA only. Bend angles are difficult to
measure. (E-F) 560 bp DNA+AraC+arabinose.
Figure 4. AFM images showing different DNA structures on mica for 560 bp
DNA. Width of image = 250 nm. (A) Simple structure showing how bend
angles were measured. The red lines are tangent to the contour on either
side of the bend. Bend angles were not measured for (B) highly curved
and (C) segmented structures. A small hook at the end of the DNA, shown
in (A), or a circular feature, shown in (B) and (C) was ignored for bend
angle measurements.
Figure 5. DNA bend angle histograms. Gaussian functions were fit to the
data, excluding 0° bend angles. Mean and standard deviation are shown.
(A) Samples prepared with DNA, AraC, and arabinose. Data for 302 bp and
560 bp were combined. Average bend angle: 69° ± 25°, N = 267; (B) 302 bp
DNA only, 47° ± 27°, N = 88; (C) 302 bp DNA, AraC, arabinose, 75° ± 26°,
N = 110; (D) 560 bp DNA, AraC, arabinose, 65° ± 21°, N = 157.
Figure 6. Initial construction of the AraC/DNA model prior to
large-scale bending of DNA in the spacer region. (A) DBD-A/I1complex is docked onto both DDs. I1 (medium gray); DBD-A (white);
DDs (dark gray). Surface representation. (B) Full I1-spacer-I2 is
shown with the DBD-A/I1 /DDs complex (left) and DBD-B/I2complex (right). Cartoon representation.
Figure 7. Full model showing the bending of DNA. N = N-terminus. (A)
Cartoon representation. Orange line segments show a bend angle of 69°.
DDs (dark gray), DBD-A (light blue), DBD-B (dark blue), DNA (gray), 4-bp
spacer region (pink). I1 is above the spacer. Teal lines,
representing the 7-residue linkers, are drawn between Cαof I167 of the DD and M175 of the DBD. (B) Surface representation
showing DNA bending. (C) Alternate view rotated by 90° from (B). Both
DDs are visible. (D) Schematic diagram corresponding to (B) indicating
the domains of AraC and I1-spacer-I2 . (E) Schematic diagram
corresponding to (C).
Figure 8. A schematic diagram approximately to scale of the sizes,
relative binding positions, and DNA bends induced by CRP, AraC, and
RNAP.
Figure S1. Full sequences of the plasmid, insert, primers, and PCR
conditions for producing the 302 bp and 560 bp DNA fragments used in
this study.
Figure S2. Histogram of measured lengths of 560 bp DNA. No AraC or
arabinose was added. A Gaussian function was used to fit the histogram
data and determine the mean and standard deviation of the length
distribution. Length = 159 ± 24 nm, N = 131.
Figure S3. PyMol session file showing the model of AraC and bent DNA.
The user can click on the DD and DBD domains of subunits A and B;araI1 , araI2 , spacer, key biochemical contacts in the
protein and DNA, and the Cα endpoints of linkers A and
B. Filename: Figure_S3_structure_AraC_DNA.pse.