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. 2018 Mar 16;46(5):2636-2647.
doi: 10.1093/nar/gky033.

Experimental maps of DNA structure at nucleotide resolution distinguish intrinsic from protein-induced DNA deformations

Robert N Azad  1 , Dana Zafiropoulos  1 , Douglas Ober  1 , Yining Jiang  1 , Tsu-Pei Chiu  2 , Jared M Sagendorf  2 , Remo Rohs  2 , Thomas D Tullius  1   3
Affiliations

Experimental maps of DNA structure at nucleotide resolution distinguish intrinsic from protein-induced DNA deformations

Robert N Azad et al. Nucleic Acids Res. .

Abstract

Recognition of DNA by proteins depends on DNA sequence and structure. Often unanswered is whether the structure of naked DNA persists in a protein-DNA complex, or whether protein binding changes DNA shape. While X-ray structures of protein-DNA complexes are numerous, the structure of naked cognate DNA is seldom available experimentally. We present here an experimental and computational analysis pipeline that uses hydroxyl radical cleavage to map, at single-nucleotide resolution, DNA minor groove width, a recognition feature widely exploited by proteins. For 11 protein-DNA complexes, we compared experimental maps of naked DNA minor groove width with minor groove width measured from X-ray co-crystal structures. Seven sites had similar minor groove widths as naked DNA and when bound to protein. For four sites, part of the DNA in the complex had the same structure as naked DNA, and part changed structure upon protein binding. We compared the experimental map with minor groove patterns of DNA predicted by two computational approaches, DNAshape and ORChID2, and found good but not perfect concordance with both. This experimental approach will be useful in mapping structures of DNA sequences for which high-resolution structural data are unavailable. This approach allows probing of protein family-dependent readout mechanisms.

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Figures

Figure 1.
Figure 1.
Comparison of the patterns of DNA minor groove width variation in naked DNA and in protein–DNA complexes. Blue, minor groove width measured from X-ray co-crystal structures of protein–DNA complexes; red, the ORChID2 pattern determined experimentally for a 399-bp DNA molecule containing 11 protein–DNA binding sites and the Drew-Dickerson dodecamer sequence (Dickerson).
Figure 2.
Figure 2.
For some protein–DNA complexes, the pattern of minor groove width variation is similar to that of the same sequence as naked DNA. Patterns were quantitatively compared by computing the Spearman's rank correlation coefficient ρ and the P-value for the similarity. The y-axis scale for expORChID2 values differs slightly between plots to facilitate comparison of individual patterns. This does not affect the calculation of the Spearman's rank correlation coefficient (see Materials and Methods). Red filled circles, expORChID2 values; blue filled triangles, minor groove width measured from the protein–DNA complex. Arrows, locations of arginine residues bound to the minor groove in the protein–DNA complex, for reference. (A) Ubx-Exd; (B) Phage 434 repressor; (C) Pit-1.
Figure 3.
Figure 3.
For some protein–DNA complexes, the pattern of minor groove width variation is similar to that of the same sequence as naked DNA. Patterns were quantitatively compared by computing the Spearman's rank correlation coefficient ρ and the P-value for the similarity. The y-axis scale for expORChID2 values differs slightly between plots to facilitate comparison of individual patterns. This does not affect the calculation of the Spearman's rank correlation coefficient (see Materials and Methods). Red filled circles, expORChID2 values; blue filled triangles, minor groove width measured from the protein–DNA complex. Arrows, locations of arginine residues bound to the minor groove in the protein–DNA complex, for reference. (A) Oct-1; (B) MogR; (C) Msx-1; (D) MATa1-MATα2.
Figure 4.
Figure 4.
For some protein–DNA complexes, the pattern of minor groove width variation in part of the binding site is similar to that of the same sequence as naked DNA, and in part of the binding site the pattern is different when protein is bound. Patterns were quantitatively compared by computing the Spearman's rank correlation coefficient ρ and the P-value for the similarity. The y-axis scale for expORChID2 values differs slightly between plots to facilitate comparison of individual patterns. This does not affect the calculation of the Spearman's rank correlation coefficient (see Materials and Methods). Red filled circles, expORChID2 values; blue filled triangles, minor groove width measured from the protein–DNA complex. Arrows, locations of arginine residues bound to the minor groove in the protein–DNA complex, for reference. (A) Tc3; (B) PhoB; (C) MATα2-MCM1; (D) Oct-1 (PORE). Dashed green lines in (D) demarcate segments of the binding site that interact with the POU-homeodomains (left and right sides) and the POU-specific domains (center) of the Oct-1 (PORE) binding site (see Supplementary Figure S6 for more details).
Figure 5.
Figure 5.
Computational prediction of minor groove width patterns of naked DNA by DNAshape sometimes differs from experimental patterns. Patterns were quantitatively compared by computing the Spearman's rank correlation coefficient ρ and the P-value for the similarity. The y-axis scale for expORChID2 values differs slightly between plots to facilitate comparison of individual patterns. This does not affect the calculation of the Spearman's rank correlation coefficient (see text). Red filled circles, expORChID2 values; teal filled squares, minor groove width predicted by DNAshape for naked DNA. (A) Oct-1 (PORE); (B) Pit-1.
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References

    1. Garvie C.W., Wolberger C.. Recognition of specific DNA sequences. Mol. Cell. 2001; 8:937–946. - PubMed
    1. Locasale J.W., Napoli A.A., Chen S., Berman H.M., Lawson C.L.. Signatures of protein–DNA recognition in free DNA binding sites. J. Mol. Biol. 2009; 386:1054–1065. - PMC - PubMed
    1. Slattery M.G., Zhou T., Yang L., Dantas Machado A.C., Gordân R., Rohs R.. Absence of a simple code: how transcription factors read the genome. Trends Biochem. Sci. 2014; 39:381–399. - PMC - PubMed
    1. Zentner G.E., Kasinathan S., Xin B., Rohs R., Henikoff S.. ChEC-seq kinetics discriminates transcription factor binding sites by DNA sequence and shape in vivo. Nat. Commun. 2015; 6:8733. - PMC - PubMed
    1. Mathelier A., Xin B., Chiu T.P., Yang L., Rohs R., Wasserman W.W.. DNA shape features improve transcription factor binding site predictions in vivo. Cell Syst. 2016; 3:278–286. - PMC - PubMed

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