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Mechanism of DNA flexibility enhancement by HMGB proteins.

Zhang J, McCauley MJ, Maher LJ, Williams MC, Israeloff NE - Nucleic Acids Res. (2009)

Bottom Line: These results are consistent with analysis of the observed global persistence length changes derived from end-to-end distance measurements, and with results of DNA-stretching experiments.The moderately broad distributions of bend angles induced by both proteins are inconsistent with either a static kink model, or a purely flexible hinge model for DNA distortion by protein binding.Therefore, the mechanism by which HMGB proteins enhance the flexibility of DNA must differ from that of the Escherichia coli HU protein, which in previous studies showed a flat angle distribution consistent with a flexible hinge model.

View Article: PubMed Central - PubMed

Affiliation: Department of Physics, Northeastern University, Boston, MA 02115, USA.

ABSTRACT
The mechanism by which sequence non-specific DNA-binding proteins enhance DNA flexibility is studied by examining complexes of double-stranded DNA with the high mobility group type B proteins HMGB2 (Box A) and HMGB1 (Box A+B) using atomic force microscopy. DNA end-to-end distances and local DNA bend angle distributions are analyzed for protein complexes deposited on a mica surface. For HMGB2 (Box A) binding we find a mean induced DNA bend angle of 78 degrees, with a standard error of 1.3 degrees and a SD of 23 degrees, while HMGB1 (Box A+B) binding gives a mean bend angle of 67 degrees, with a standard error of 1.3 degrees and a SD of 21 degrees. These results are consistent with analysis of the observed global persistence length changes derived from end-to-end distance measurements, and with results of DNA-stretching experiments. The moderately broad distributions of bend angles induced by both proteins are inconsistent with either a static kink model, or a purely flexible hinge model for DNA distortion by protein binding. Therefore, the mechanism by which HMGB proteins enhance the flexibility of DNA must differ from that of the Escherichia coli HU protein, which in previous studies showed a flat angle distribution consistent with a flexible hinge model.

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(a) The distribution of bare dsDNA contour lengths and its Gaussian fit. (b) The local bend angle distribution for bare dsDNA (segment step size 10 nm) and Gaussian fit.
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Figure 2: (a) The distribution of bare dsDNA contour lengths and its Gaussian fit. (b) The local bend angle distribution for bare dsDNA (segment step size 10 nm) and Gaussian fit.

Mentions: Bare linearized dsDNA from plasmid pBR322 (4361 bp) was initially imaged (Figure 1). The distribution of contour lengths obtained from 40 molecules by image tracing (Figure 2a) had a mean of 1.48 μm, and SD σ = 0.11 μm, giving 0.34 ± 0.03 nm per bp, which agrees very well with the theoretical value of 0.34 nm per bp in B-form dsDNA (28). The local DNA bend angle is defined as the angle between the pair of adjacent line segments formed with three neighboring points along the contour, as shown in Figure 1b. Bend angles were measured for points at 10 nm intervals on the bare DNA molecules. The distribution of local bend angles was Gaussian-like (29), with a mean of 0° and σ ∼22° (Figure 2b). However, resolution effects give an apparent width of DNA strands of approximately 15 nm, which causes measurement error in the angle data, particularly for small segment sizes. The image tracing algorithm affects this error somewhat, hence the error was estimated from the angle variance in visually straight sections of DNA. We estimated the angle error to be σ′ ∼ 8° for 10 nm segments. Therefore, the intrinsic DNA bend angle distribution should have width: ∼20.5°. For smaller segment sizes the angle error increases, as will be shown in the analysis of persistence length discussed below.Figure 1.


Mechanism of DNA flexibility enhancement by HMGB proteins.

Zhang J, McCauley MJ, Maher LJ, Williams MC, Israeloff NE - Nucleic Acids Res. (2009)

(a) The distribution of bare dsDNA contour lengths and its Gaussian fit. (b) The local bend angle distribution for bare dsDNA (segment step size 10 nm) and Gaussian fit.
© Copyright Policy - creative-commons
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2651801&req=5

Figure 2: (a) The distribution of bare dsDNA contour lengths and its Gaussian fit. (b) The local bend angle distribution for bare dsDNA (segment step size 10 nm) and Gaussian fit.
Mentions: Bare linearized dsDNA from plasmid pBR322 (4361 bp) was initially imaged (Figure 1). The distribution of contour lengths obtained from 40 molecules by image tracing (Figure 2a) had a mean of 1.48 μm, and SD σ = 0.11 μm, giving 0.34 ± 0.03 nm per bp, which agrees very well with the theoretical value of 0.34 nm per bp in B-form dsDNA (28). The local DNA bend angle is defined as the angle between the pair of adjacent line segments formed with three neighboring points along the contour, as shown in Figure 1b. Bend angles were measured for points at 10 nm intervals on the bare DNA molecules. The distribution of local bend angles was Gaussian-like (29), with a mean of 0° and σ ∼22° (Figure 2b). However, resolution effects give an apparent width of DNA strands of approximately 15 nm, which causes measurement error in the angle data, particularly for small segment sizes. The image tracing algorithm affects this error somewhat, hence the error was estimated from the angle variance in visually straight sections of DNA. We estimated the angle error to be σ′ ∼ 8° for 10 nm segments. Therefore, the intrinsic DNA bend angle distribution should have width: ∼20.5°. For smaller segment sizes the angle error increases, as will be shown in the analysis of persistence length discussed below.Figure 1.

Bottom Line: These results are consistent with analysis of the observed global persistence length changes derived from end-to-end distance measurements, and with results of DNA-stretching experiments.The moderately broad distributions of bend angles induced by both proteins are inconsistent with either a static kink model, or a purely flexible hinge model for DNA distortion by protein binding.Therefore, the mechanism by which HMGB proteins enhance the flexibility of DNA must differ from that of the Escherichia coli HU protein, which in previous studies showed a flat angle distribution consistent with a flexible hinge model.

View Article: PubMed Central - PubMed

Affiliation: Department of Physics, Northeastern University, Boston, MA 02115, USA.

ABSTRACT
The mechanism by which sequence non-specific DNA-binding proteins enhance DNA flexibility is studied by examining complexes of double-stranded DNA with the high mobility group type B proteins HMGB2 (Box A) and HMGB1 (Box A+B) using atomic force microscopy. DNA end-to-end distances and local DNA bend angle distributions are analyzed for protein complexes deposited on a mica surface. For HMGB2 (Box A) binding we find a mean induced DNA bend angle of 78 degrees, with a standard error of 1.3 degrees and a SD of 23 degrees, while HMGB1 (Box A+B) binding gives a mean bend angle of 67 degrees, with a standard error of 1.3 degrees and a SD of 21 degrees. These results are consistent with analysis of the observed global persistence length changes derived from end-to-end distance measurements, and with results of DNA-stretching experiments. The moderately broad distributions of bend angles induced by both proteins are inconsistent with either a static kink model, or a purely flexible hinge model for DNA distortion by protein binding. Therefore, the mechanism by which HMGB proteins enhance the flexibility of DNA must differ from that of the Escherichia coli HU protein, which in previous studies showed a flat angle distribution consistent with a flexible hinge model.

Show MeSH
Related in: MedlinePlus