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The structure of DNA by direct imaging.

Marini M, Falqui A, Moretti M, Limongi T, Allione M, Genovese A, Lopatin S, Tirinato L, Das G, Torre B, Giugni A, Gentile F, Candeloro P, Di Fabrizio E - Sci Adv (2015)

Bottom Line: In the image, all relevant lengths of A-form DNA are measurable.A high-resolution transmission electron microscope that operates at 80 keV with an ultimate resolution of 1.5 Å was used for this experiment.Direct imaging of a single molecule can be used as a method to address biological problems that require knowledge at the single-molecule level, given that the average information obtained by x-ray diffraction of crystals or fibers is not sufficient for detailed structure determination, or when crystals cannot be obtained from biological molecules or are not sufficient in understanding multiple protein configurations.

View Article: PubMed Central - PubMed

Affiliation: SMILEs Lab, Physical Science and Engineering (PSE) and Biological and Environmental Science and Engineering (BESE) Divisions, King Abdullah University of Science and Technology, Thuwal 23955-6900, Kingdom of Saudi Arabia.

ABSTRACT
The structure of DNA was determined in 1953 by x-ray fiber diffraction. Several attempts have been made to obtain a direct image of DNA with alternative techniques. The direct image is intended to allow a quantitative evaluation of all relevant characteristic lengths present in a molecule. A direct image of DNA, which is different from diffraction in the reciprocal space, is difficult to obtain for two main reasons: the intrinsic very low contrast of the elements that form the molecule and the difficulty of preparing the sample while preserving its pristine shape and size. We show that through a preparation procedure compatible with the DNA physiological conditions, a direct image of a single suspended DNA molecule can be obtained. In the image, all relevant lengths of A-form DNA are measurable. A high-resolution transmission electron microscope that operates at 80 keV with an ultimate resolution of 1.5 Å was used for this experiment. Direct imaging of a single molecule can be used as a method to address biological problems that require knowledge at the single-molecule level, given that the average information obtained by x-ray diffraction of crystals or fibers is not sufficient for detailed structure determination, or when crystals cannot be obtained from biological molecules or are not sufficient in understanding multiple protein configurations.

No MeSH data available.


Related in: MedlinePlus

Base propeller twist.A representative image of simulating different amounts of propeller twist of the A-T couple of bases in A-DNA is shown.
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Figure 2: Base propeller twist.A representative image of simulating different amounts of propeller twist of the A-T couple of bases in A-DNA is shown.

Mentions: The last important quantitative information we can obtain from the TEM image is the orientation of the helix. From Fig. 1D, in particular, we measure the angle α between the helix axis and the normal to the base pair, being α = 19°. This is the distinctive spatial configuration of dsDNA when humidity drops below 75% (9, 14, 15). All these values are summarized in Table 1; in the third column, we report the x-ray accepted data and modeling values for comparison with our direct imaging. We notice that five new molecular structures are measured in the present study. Finally, we notice from Fig. 1 that the variable intensity contrast in the inner of the helix, which we attribute to the base pairs, is due to the positive propeller twist (13) of the bases with respect to the horizontal plane orthogonal to the DNA axis, as explicitly shown in Fig. 2. We also point out that in Table 1, the difference in length between the DNA diameter and the sum of the lengths of the base pairs and the backbone can be attributed to hydrogen bonds, which are between 2 and 3 Å, depending on the stretching state of the nucleotide in the helix position.


The structure of DNA by direct imaging.

Marini M, Falqui A, Moretti M, Limongi T, Allione M, Genovese A, Lopatin S, Tirinato L, Das G, Torre B, Giugni A, Gentile F, Candeloro P, Di Fabrizio E - Sci Adv (2015)

Base propeller twist.A representative image of simulating different amounts of propeller twist of the A-T couple of bases in A-DNA is shown.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4643809&req=5

Figure 2: Base propeller twist.A representative image of simulating different amounts of propeller twist of the A-T couple of bases in A-DNA is shown.
Mentions: The last important quantitative information we can obtain from the TEM image is the orientation of the helix. From Fig. 1D, in particular, we measure the angle α between the helix axis and the normal to the base pair, being α = 19°. This is the distinctive spatial configuration of dsDNA when humidity drops below 75% (9, 14, 15). All these values are summarized in Table 1; in the third column, we report the x-ray accepted data and modeling values for comparison with our direct imaging. We notice that five new molecular structures are measured in the present study. Finally, we notice from Fig. 1 that the variable intensity contrast in the inner of the helix, which we attribute to the base pairs, is due to the positive propeller twist (13) of the bases with respect to the horizontal plane orthogonal to the DNA axis, as explicitly shown in Fig. 2. We also point out that in Table 1, the difference in length between the DNA diameter and the sum of the lengths of the base pairs and the backbone can be attributed to hydrogen bonds, which are between 2 and 3 Å, depending on the stretching state of the nucleotide in the helix position.

Bottom Line: In the image, all relevant lengths of A-form DNA are measurable.A high-resolution transmission electron microscope that operates at 80 keV with an ultimate resolution of 1.5 Å was used for this experiment.Direct imaging of a single molecule can be used as a method to address biological problems that require knowledge at the single-molecule level, given that the average information obtained by x-ray diffraction of crystals or fibers is not sufficient for detailed structure determination, or when crystals cannot be obtained from biological molecules or are not sufficient in understanding multiple protein configurations.

View Article: PubMed Central - PubMed

Affiliation: SMILEs Lab, Physical Science and Engineering (PSE) and Biological and Environmental Science and Engineering (BESE) Divisions, King Abdullah University of Science and Technology, Thuwal 23955-6900, Kingdom of Saudi Arabia.

ABSTRACT
The structure of DNA was determined in 1953 by x-ray fiber diffraction. Several attempts have been made to obtain a direct image of DNA with alternative techniques. The direct image is intended to allow a quantitative evaluation of all relevant characteristic lengths present in a molecule. A direct image of DNA, which is different from diffraction in the reciprocal space, is difficult to obtain for two main reasons: the intrinsic very low contrast of the elements that form the molecule and the difficulty of preparing the sample while preserving its pristine shape and size. We show that through a preparation procedure compatible with the DNA physiological conditions, a direct image of a single suspended DNA molecule can be obtained. In the image, all relevant lengths of A-form DNA are measurable. A high-resolution transmission electron microscope that operates at 80 keV with an ultimate resolution of 1.5 Å was used for this experiment. Direct imaging of a single molecule can be used as a method to address biological problems that require knowledge at the single-molecule level, given that the average information obtained by x-ray diffraction of crystals or fibers is not sufficient for detailed structure determination, or when crystals cannot be obtained from biological molecules or are not sufficient in understanding multiple protein configurations.

No MeSH data available.


Related in: MedlinePlus