Organic Pedagogical Electronic Network

Cyclic Conformation and Nucleic Acid Sugar Puckering

Roland Jones, Dane Brankle, and Peter StevensonUniversity of Utah

https://commons.wikimedia.org/wiki/File:A-B-Z-DNA_Side_View.png

Conformation of cyclic systems and illustration

Ansyln, E.V.; Dougherty, D.A. Modern Physical Organic Chemistry. University Science Books, 2006. Saenger, W. Principles of Nucleic Acid Structure, Springer-Verlag, 1984. http://x3dna.org/highlights/sugar-pucker-correlates-with-phosphorus-base-distance. http://casegroup.rutgers.edu/lnotes/dnab.pdf.

Model cyclic systems: Cyclic systems are ubiquitous. Establishing an understanding of the shape preferences (e.g., strain and energetics) regarding representative cyclic models is a powerful tool in conformational analysis. The expanded review of fundamental cycloalkanes can further assist in preferential conformational analysis of associated derivatives.

Much of the resulting conformational preferences can be attributed to bond angle strain and/or C-H interactions (e.g., torsional strain). Some general systems are depicted here. (1) Cyclopropane is planar (as a result of the carbon framework) with a strain energy of 27.5 kcal/mol. (2) Cyclobutane is not planar and assumes a puckered conformation to relieve C-H eclipsing. It has a strain energy of 26.5 kcal/mol. (3) Cyclopentane is also puckered to relieve C-H eclipsing. It has a strain energy of 6.2 kcal/mol. Finally, (4) cyclohexane predominantly assumes a rigid chair conformation providing a commonly assigned strain energy of 0 kcal/mol.

(1) Cycloproane (2) Cyclobutane (3) Cyclopentane (4) Cyclohexane

Conformation of cyclic systems and illustration

Ansyln, E.V.; Dougherty, D.A. Modern Physical Organic Chemistry. University Science Books, 2006. Saenger, W. Principles of Nucleic Acid Structure, Springer-Verlag, 1984. http://x3dna.org/highlights/sugar-pucker-correlates-with-phosphorus-base-distance. http://casegroup.rutgers.edu/lnotes/dnab.pdf.

Illustrative Effect: Resulting conformational dynamics can easily be illustrated upon review of the biological ramifications concerning cyclopentane in DNA conformations. The favored conformation of DNA building blocks plays a crucial role in defining the resulting structure (or global conformation) of DNA. As discussed above (i.e., a five-membered ring), deoxyribose sugar rings contain five atoms that cannot lie in one plane: at least one must be out of plane. It can then be anticipated that the alternating conformations of deoxyribose rings influence overall DNA conformation.

Conformation: There are two predominate deoxyribose ring conformations that strongly influence the global conformation of DNA: C3’-endo and C2’-endo sugar puckers.

C2’-endo is an S-type conformation(S for south)

C3’-endo is an N-type conformation(N for north)

Nucleic acid sugar puckering

History: Over 60 years ago, analysis of the diffraction pattern of DNA was consistent with a double helix conformation. However, what was not immediately answered was why there were various forms of DNA observed: predominately A-form and B-form. Over the subsequent years, the resulting conformation of the deoxyribose sugar ring provided insight into the global conformation of DNA.

Saenger, W. Principles of Nucleic Acid Structure, Springer-Verlag, 1984. Rich, A. Nat. Struct. Bio. 2003. 10, 247-249. Patel, D.; et al. Proc. Natl. Acad. Sci. 1978. 75, 6, 2533-2557. Williams, A.A.: et al. Biochemistry. 2009. 48, 11994-12004. https://commons.wikimedia.org/wiki/File:A-B-Z-DNA_Side_View.png.

A-form B-form

Puckering Impact: With continued analysis, it became apparent that the B-form contains a ring pucker in which the C2’ atom is out of plane on the same side as the base (C2’-endo).

Consequently, the phosphate groups are about 7 Å apart. Subsequent evaluation of the A-form determined that the C3’ atom is out of plane (C3’-endo). The resulting conformation positions the phosphate groups at about 5.9 Å apart. Consequently, the sugar phosphate backbone is shorter in comparison to the C2’-endo conformation, resulting in a double helix in which the base pairs are somewhat displaced.

Nucleic acid sugar puckering

Saenger, W. Principles of Nucleic Acid Structure, Springer-Verlag, 1984. Rich, A. Nat. Struct. Bio. 2003. 10, 247-249. Patel, D.; et al. Proc. Natl. Acad. Sci. 1978. 75, 6, 2533-2557. Williams, A.A.: et al. Biochemistry. 2009. 48, 11994-12004. https://commons.wikimedia.org/wiki/File:A-B-Z-DNA_Side_View.png.

Z-form

Left-Handed DNA: As the conformational impacts were understood, additional efforts led to the evaluation of an unusual left-handed double helix. Still held together with Watson-Crick base pairs (i.e., d(CGCGCG)), it is somewhat elongated and thinner with only one grove in comparison to A- and B-forms. It is related to B-form by flipping the bases upside down so that the upper surface of a base pair in B-DNA becomes the lower surface of a the base pair when it changes to Z-DNA. This inversion results in rotating the purine bases about their C-N glycosyl bond (depicted in red below), so that it assumes the syn conformation.

NH

N

N

O

NH2N

O

HOH

HHHH

HO

Anti

HN

N

N

O

H2N N

O

HOH

HHHH

HO

Syn

Supplementary and illustrative studies

Moreau, C.; et al. Org. Biomol. Chem. 2011. 1, 278-290.

Case Study I

DNA sugar pucker is sensitive to many substituents that are placed on the deoxyribose nitrogenous base. A common substituent placed on the base Is a methyl group on the C5’ carbon. This is often done naturally by the cell as an epigenetic marker for many cellular processes in both, prokaryotes and eukaryotes. Studies show that the addition of the methyl group on the cytosine as shown below causes increased contribution of C2’-endo pucker.

Supplementary and illustrative studies

Marquez, V.E; et al. J. Am. Chem. Soc. 2004. 126, 2, 543-549

Case Study II

A kinase is an enzyme that phosphorylates a certain molecule (such as the phosphorylation of the 5’ hydroxyl group depicted here). Shown below is the pseudorotation conformation cycle of a of an unrestricted furanose ring (defined by the value of P which depends on the associated torsional angles (n0 to n4)). Crystallographic data indicates that puckering preference is between the 3’-endo and 2’-endo conformations.

Supplementary and illustrative studies

Santos, R.A; et al. Biochemistry. 1989. 28, 9372-9378. Banway, M.; et al. J. Mol. Biol. 2002. 324, 667-676. Wu, Z.; et al. J. Biomol. NMR. 2003. 26, 297-315

Case Study III

Solid-state NMR is a spectroscopy method that employs assessment of anisotropic (directionally specific) interactions. These interactions modify spin energy levels for all sites in a molecule (which changes the resonance frequency).

Solid-state 13C NMR has been used with a series of crystalline nucleosides and nucleotides. Characterization provided the measurement of dexoyribose ring conformation relative to the carbon chemical shift. It was observed that 3’-endo conformers have 3’ and 5’ chemical shifts more upfield in comparison to 2’-endo conformers.

DNA C1’ C2’ C3’ C4’ C5’

A-form 81.6 38.7 68.2 83.6 60.9

B-form 84.4 36.3 77.3 84.4 66.2

Average chemical shift in ppm for two principle forms of DNA.

Problems1) Rank the illustrated representative cycloalkane molecules in order from most to least stable and most to least acidic.

2) If a double bond were added to each of the illustrated representative cycloalkane molecules, how would the resulting strain energies be affected? Why?

3) Draw the most likely conformation of the deoxyribonucleoside shown here.

4) Propose an explanation for why methylation of cytosine causes an increase in the contribution of C2’-endo pucker versus C3’-endo pucker (see Case Study I).

5) Predict the preferred puckered conformation of the 5’ hydroxyl ring shown below in the presence of a catalyzing kinase enzyme under characteristic reaction conditions (see Case Study II).

6) Describe how 13C NMR can be used as a probe for DNA ring puckering. Suggest alternative probes in characterizing and analyzing sugar ring puckering (see Case Study III and references).

NH

N

N

O

NH2N

O

HOH

HHHH

HO

OBHO

Solutions1) Stability: cyclohexane > cyclopentane > cyclobutane > cyclopropane. Acidity: cyclopropane > cyclobutane > cyclopropane >

cyclohexane.

2) Incorporating an olefin into a small ring generally increases the strain energy. Olefins prefer larger angles over alkanes resulting in more destabilized cyclic systems (with a more noticeable effect seen in cyclopropene, cyclobutane, and cyclohexane). However, the strain energy is lower in cyclopentene as it has only one set of C-H bonds eclipsed versus two sets in cyclopentane. The preferred (rigid) chair conformation is disrupted in cyclohexane with the addition of an olefin resulting in a slight strain energy increase (about 0.4 kcal/mol).

3) In five membered rings, the anomeric effect is less pronounced. Consequently, the purine substituent will be in more of an equatorial position with either C2’- or C3’-endo conformation upon puckering. The least torsional strain is seen when the purine (or base) substituent is in the anti orientation. Rotation about the C-N glycosyl bond will contribute to modifications in resulting DNA conformation (such as Z-DNA).

4) As discussed in the overview, a C3’-endo pucker causes the sugar backbone to shorten compared a C2’-endo pucker. This also causes the base substituents to be slightly displaced. Therefore, a bigger base (such as methylated cytosine) prefers more space due to steric repulsions.

5) Studies show that kinase activity often prefers the C2’-endo (south) pucker because the hydroxyl group is more easily accessed through a sense of steric activity.

6) The 13C chemical shift will vary relative to the puckering conformation. Additionally, functionalization could be evaluated via FTIR (such as DNA methylation which has significance in transcription). Depending on synthetic difficulties, isotopic labeling could be employed relative to the desired assessment (e.g., conformation, mechanism, etc.).

See previously cited references for further details and review.

H

HH

H

HH

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Contributed by: Roland Jones, Dane Brankle, and Peter Stevenson

University of Utah

2015