In Glen Report 33.1 we discussed the standard numbering system for nucleobases in DNA and RNA nucleotide building blocks.1 In this note, we would like to take a closer look at another necessary component in the nucleotide: the sugar ring. One of the few structural differences between DNA and RNA is the lack of a 2’-hydroxyl group in the ribose ring of DNA. This seemingly minor change in the structure has significant biological consequences.
Conformation and dynamics of the (deoxy)ribose impact the overall structure of the nucleic acid, which contributes to recognition and function of the genomic material. The presence of a 2’- functional group influences the more favorable sugar pucker conformation.2 In turn, the sugar pucker defines the predominant structure adopted upon base pairing interactions (Figure 1). Four conformations are possible: C3’-endo, C2’-endo, O4’-exo and O4’-endo.
Figure 1. Sugar Pucker Conformations
DNA adopts a C2’-endo sugar pucker (also referred to as the South conformer), which corresponds to the favored B-form of DNA, where base pairs are almost centered over the helical axis (Figure 2A). In contrast, RNA consists of the C3’-endo sugar pucker (North conformer) due to the size of the C2’-substitution (Figure 2A). As a result, the nucleic acid adopts an A-form, where base pairs are displaced away from the central axis and closer to the major groove. A-form RNA resembles a ribbon-like helix with an open center whereas B-form DNA is the canonical double helix.
Figure 2. Sugar Conformers
Recently, more sugar modifications have been described and studied, especially due to their relevancy in oligonucleotide therapeutics. For this article, we will focus on the sugar modifications that we offer: Locked Analog (LA), 2’-OMe, 2’-MOE, 2’-F RNA, and 2’-F-ANA (Figure 3).
Figure 3. Sugar modifications available at Glen Research
We have previously reported on the benefits of our locked analog series.3,4 Locked nucleic acids offer enhanced binding and allow controlled manipulation of melting temperatures. Our LA modifications are often used in hybridization probes but are also becoming more popular in antisense oligonucleotide therapy. LNA modifications reinforce the C3’-endo conformation. This backbone is so popular that we recently introduced the LA solid supports to accompany our LA phosphoramidites.4
Our 2’-alkoxy modifications include 2’-OMe and 2’-MOE. Nucleic acids bearing a 2’-OMe occur naturally in tRNA and other post-translationally modified small RNAs. The same cannot be said for 2’-MOE. The substitution of 2’-hydroxyl to 2’-O-alkyl results in increased chemical and duplex stability and nuclease resistance. The low toxicity of these modifications makes them attractive backbones for therapeutic candidates. Duplexes with 2’-OMe or 2’-MOE may give rise to additional stabilizing forces from base stacking and hydrophobic interactions in the minor groove.2 However, the 2’-O-alkyl substituents have not been shown to enhance base pairing.
Depending on the orientation of the substituent, the C2’-F group will determine the sugar conformation due to its polarity, rather than its size. The high electronegativity of the fluoride draws the C2’-bonds towards it.2 Due to these interactions, 2’-F-RNA modifications adopt the standard C3’-endo pucker. Conversely, 2’-F-ANA adopt a B-form helix, although it is through an O4’-endo pucker (Figure 2B). The 2’-F substitutions provide enhanced thermal stability and binding effects. Out of the two, only the 2’-F arabinose substituents are resistant to nucleases. Modifications with fluorine substituents can be monitored by NMR.
Overall, modifications on the furanose ring can yield subtle to major effects on the structure and function of the nucleic acid. Properties such as resistance to nucleases, enhanced chemical and thermal stability, and the strength of base pairing interactions, can easily be fine-tuned to best fit one’s application needs (Table 1).
Table 1. Overview of standard and modified sugar backbones available at Glen Research
|
Backbone |
Sugar Pucker |
Form |
Nuclease Resistance |
Thermal Stability |
Provides Stronger Base Pairing |
|
DNA |
C2’-endo |
B |
|
|
|
|
RNA |
C3’-endo |
A |
|
|
|
|
LA |
C3’-endo |
A |
✓ |
✓ |
✓ |
|
2’-OMe |
C3’-endo |
A |
✓ |
✓ |
|
|
2’-MOE |
C3’-endo |
A |
✓ |
✓ |
|
|
2’-F |
C3’-endo |
A |
|
✓ |
✓ |
|
2’-F-ANA |
O4’-endo |
B |
✓ |
✓ |
✓ |
References
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