Does Inter-Residue Hydrogen Bonding in ß-(1→4)-Linked Disaccharides Influence Linkage Conformation in Aqueous Solution?

Two disaccharides, methyl ß-d-galactopyranosyl-(1→4)-α-d-glucopyranoside (1) and methyl ß-d-galactopyranosyl-(1→4)-3-deoxy-α-d-ribo-hexopyranoside (3), were prepared with selective 13C-enrichment to allow measurement of six trans-O-glycosidic J-couplings (2JCOC, 3JCOCH, and 3JCOCC) in each compound. Density functional theory (DFT) was used to parameterize Karplus-like equations that relate these J-couplings to either ϕ or ψ. MA'AT analysis was applied to both linkages to determine mean values of ϕ and ψ in each disaccharide and their associated circular standard deviations (CSDs). Results show that deoxygenation at C3 of 1 has little effect on both the mean values and librational motions of the linkage torsion angles. This finding implies that, if inter-residue hydrogen bonding between O3H and O5' of 1 is present in aqueous solution and persistent, it plays little if any role in dictating preferred linkage conformation. Hydrogen bonding may lower the energy of the preferred linkage geometry but does not determine it to any appreciable extent. Aqueous 1-µs MD simulation supports this conclusion and also indicates greater conformational flexibility in deoxydisaccharide 3 in terms of sampling several, conformationally distinct, higher-energy conformers in solution. The populations of these latter conformers are low (3-14%) and could not be validated by MA'AT analysis. If the MD model is correct, however, C3 deoxygenation does enable conformational sampling over a wider range of ϕ/ψ values, but linkage conformation in the predominant conformer is essentially identical in both 1 and 3.

Methods of study on conformation of polysaccharides from natural products: A review.

Polysaccharides from natural products play multiple roles and have extensive bioactivities in life process. Bioactivities of polysaccharides (e.g., Lentinan, Schizophyllan, Scleroglucan, Curdlan, Cinerean) have a close relation to their chain conformation. Compared to other types of polysaccharides, the conformation of ß-glucan has been studied more. The major research methods of conformation of polysaccharides from natural products (Congo red experiment, circular dichroism spectrum, viscosity method, light scattering method, size exclusion chromatography, atomic force microscope), corresponding experimental schemes, and the external factors affecting polysaccharide conformation were reviewed in this paper. These research methods of conformation have been widely used, among which Congo red experiment and viscosity method are the most convenient ones to study the morphological changes of polysaccharide chains.

Revealing the key structural features promoting the helical conformation in algal polysaccharide carrageenan in solution.

Carrageenans are industrially important polysaccharides with tunable viscoelastic and gelation properties. The function of polysaccharide depends on its conformation and chemical composition. However, the solution conformations of carrageenans are highly debated, and the structure-function relationship remains elusive. Here, we have studied the intrinsic conformational behavior of a series of carrageenan hexamers in solution, using extensive all-atom classical MD and enhanced sampling. Our findings comprehensively delineate that carrageenans containing the 3,6-anhydrous bridge (κ-C, ι-C, θ-C, and non-sulfated ß-C) adopt compact helical structures as their predominant conformation in solution, whereas carrageenans without the bridge (µ-C, ν-C, and λ-C) remain as extended loosely packed helices; opposing the 'coil-to-helix' paradigm. Glycosidic linkages access a few allowed orientations. We hypothesize that the 3,6-anhydrous bridge, irrespective of carrageenan's sulfation pattern, is essential for stabilizing the helical conformation at the single-chain level. It provides necessary flexibility to the glycosidic linkage to sample conformations close to the experimentally derived helical structure and also prevents the sugar ring flipping. Sulfate groups mainly modify the chain stiffness due to steric and stereo-electronic effects and participate in hydrogen bonding. Such atomistic insights will be helpful for understanding the differential gelation mechanisms of carrageenans and fine-tuning polysaccharide backbone for various industrial applications.

Conformational Preferences at the Glycosidic Linkage of Saccharides in Solution as Deduced from NMR Experiments and MD Simulations: Comparison to Crystal Structures.

Glycans are central to information content and regulation in biological systems. These carbohydrate molecules are active either as oligo- or polysaccharides, often in the form of glycoconjugates. The monosaccharide entities are joined by glycosidic linkages and stereochemical arrangements are of utmost importance in determining conformation and flexibility of saccharides. The conformational preferences and population distributions at the glycosidic torsion angles φ and ψ have been investigated for O-methyl glycosides of three disaccharides where the substitution takes place at a secondary alcohol, viz., in α-l-Fucp-(1→3)-ß-d-Glcp-OMe, α-l-Fucp-(1→3)-α-d-Galp-OMe and α-d-Glcp-(1→4)-α-d-Galp-OMe, corresponding to disaccharide structural elements present in bacterial polysaccharides. Stereochemical differences at or adjacent to the glycosidic linkage were explored by solution state NMR spectroscopy using one-dimensional 1 H,1 H-NOESY NMR experiments to obtain transglycosidic proton-proton distances and one- and two-dimensional heteronuclear NMR experiments to obtain 3 JCH transglycosidic coupling constants related to torsion angles φ and ψ. Computed effective proton-proton distances from molecular dynamics (MD) simulations showed excellent agreement to experimentally derived distances for the α-(1→3)-linked disaccharides and revealed that for the bimodal distribution at the ψ torsion angle for the α-(1→4)-linked disaccharide experiment and simulation were at variance with each other, calling for further force field developments. The MD simulations disclosed a highly intricate inter-residue hydrogen bonding pattern for the α-(1→4)-linked disaccharide, including a nonconventional hydrogen bond between H5' in the glucosyl residue and O3 in the galactosyl residue, supported by a large downfield 1 H NMR chemical shift displacement compared to α-d-Glcp-OMe. Comparison of population distributions of the glycosidic torsion angles φ and ψ in the disaccharide entities to those of corresponding crystal structures highlighted the potential importance of solvation on the preferred conformation.

Synthesis and study of the glycosyl-donor properties of 2-(2,2,2-trichloroethoxy)-(3,4,6-tri-O-acetyl-1,2-dideoxy-α-d-galactopyrano)-[2,1-d]-2-oxazoline.

A synthesis of 2-(2,2,2-trichloroethoxy)-(3,4,6-tri-O-acetyl-1,2-dideoxy-α-d-galactopyrano)-[2,1-d]-2-oxazoline - a previously unknown 2-alkoxy glyco-[2,1-d]-2-oxazoline derivative with d-galacto configuration was carried out. Glycosylating activity of the obtained galactooxazoline has been studied and it has been shown that in the presence of a weak protic acid, such as sym-collidinium triflate, this substance exhibits properties of a reactive and 1,2-trans-stereoselective glycosyl donor. The homopolymerization reaction of oxazoline derivatives of sugars has been found to proceed under the same conditions, leading to the formation of pseudo-oligosaccharide products. It has been found that this undesirable side reaction could be suppressed by changing the acid catalyst concentration, resulting in the development of efficient methods for the synthesis of glycoside and oligosaccharide derivatives of ß-d-galactosamine using the synthesized 2-(2,2,2-trichloroethoxy)-2-oxazoline glycosyl donor under very mild conditions.

Adaptation of Zemplén's conditions for a simple and highly selective approach to methyl 1,2-trans glycosides.

1,2-trans methyl glycosides can be readily obtained from peracetylated sugars through their initial conversion into glycosyl iodide donors and subsequent exposure of these latter to a slight excess of sodium methoxide in methanol. Under these conditions a varied set of mono- and disaccharide precursors afforded the corresponding 1,2-trans glycosides with concomitant de-O-acetylation in satisfying yields (in the range 59-81%). A similar approach also proved effective when using GlcNAc glycosyl chloride as the donor.

Analysis and validation of overall N-glycan conformation in Privateer.

The oligosaccharides in N-glycosylation provide key structural and functional contributions to a glycoprotein. These contributions are dependent on the composition and overall conformation of the glycans. The Privateer software allows structural biologists to evaluate and improve the atomic structures of carbohydrates, including N-glycans; this software has recently been extended to check glycan composition through the use of glycomics data. Here, a broadening of the scope of the software to analyse and validate the overall conformation of N-glycans is presented, focusing on a newly compiled set of glycosidic linkage torsional preferences harvested from a curated set of glycoprotein models.

Glycosidase mechanisms: Sugar conformations and reactivity in endo- and exo-acting enzymes.

The enzymatic breakdown of carbohydrates plays a critical role in several biological events and enables the development of sustainable processes to obtain bioproducts and biofuels. In this scenario, the design of efficient inhibitors for glycosidases that can act as drug targets and the engineering of carbohydrate-active enzymes with tailored catalytic properties is of remarkable importance. To guide rational approaches, it is necessary to elucidate enzyme molecular mechanisms, in particular understanding how the microenvironment modulates the conformational space explored by the substrate. Computer simulations, especially those based on ab initio methods, have provided a suitable atomic description of carbohydrate conformations and catalytic reactions in several glycosidase families. In this review, we will focus on how the active-site topology (pocket or cleft) and mode of cleavage (endo or exo) can affect the catalytic mechanisms adopted by glycosidases, in particular the substrate conformations along the reaction coordinate.

Synthesis of fluoro- and seleno-containing D-lactose and D-galactose analogues.

Synthetic deoxy-fluoro-carbohydrate derivatives and seleno-sugars are useful tools in protein-carbohydrate interaction studies using nuclear magnetic resonance spectroscopy because of the presence of the 19F and 77Se reporter nuclei. Seven saccharides containing both these atoms have been synthesized, three monosaccharides, methyl 6-deoxy-6-fluoro-1-seleno-ß-D-galactopyranoside (1) and methyl 2-deoxy-2-fluoro-1-seleno-α/ß-D-galactopyranoside (2α and 2ß), and four disaccharides, methyl 4-O-(ß-D-galactopyranosyl)-2-deoxy-2-fluoro-1-seleno-ß-D-glucopyranoside (3), methyl 4-Se-(ß-D-galactopyranosyl)-2-deoxy-2-fluoro-4-seleno-ß-D-glucopyranoside (4), and methyl 4-Se-(2-deoxy-2-fluoro-α/ß-D-galactopyranosyl)-4-seleno-ß-D-glucopyranoside (5α and 5ß), the three latter compounds with an interglycosidic selenium atom. Selenoglycosides 1 and 3 were obtained from the corresponding bromo sugar by treatment with dimethyl selenide and a reducing agent, while compounds 2α/2ß, 4, and 5α/5ß were synthesized by the coupling of a D-galactosyl selenolate, obtained in situ from the corresponding isoselenouronium salt, with either methyl iodide or a 4-O-trifluoromethanesulfonyl D-galactosyl moiety. While benzyl ether protecting groups were found to be incompatible with the selenide linkage during deprotection, a change to acetyl esters afforded 4 in a 17% overall yield and over 9 steps from peracetylated D-galactosyl bromide. The synthesis of 5 was performed similarly, but the 2-fluoro substituent led to reduced stereoselectivity in the formation of the isoselenouronium salt (α/ß âˆ¼ 1 : 2.3). However, the ß-anomer of the uronium salt could be obtained almost pure (∼98%) by precipitation from the reaction mixture. The following displacement reaction occurred without anomerisation, affording, after deacetylation, pure 5ß.

Martini 3 Coarse-Grained Force Field for Carbohydrates.

The Martini 3 force field is a full reparametrization of the Martini coarse-grained model for biomolecular simulations. Due to the improved interaction balance, it allows for a more accurate description of condensed phase systems. In the present work, we develop a consistent strategy to parametrize carbohydrate molecules accurately within the framework of Martini 3. In particular, we develop a canonical mapping scheme which decomposes arbitrarily large carbohydrates into a limited number of fragments. Bead types for these fragments have been assigned by matching physicochemical properties of mono- and disaccharides. In addition, guidelines for assigning bonds, angles, and dihedrals were developed. These guidelines enable a more accurate description of carbohydrate conformations than in the Martini 2 force field. We show that models obtained with this approach are able to accurately reproduce osmotic pressures of carbohydrate water solutions. Furthermore, we provide evidence that the model differentiates correctly the solubility of the polyglucoses dextran (water-soluble) and cellulose (water insoluble but soluble in ionic liquids). Finally, we demonstrate that the new building blocks can be applied to glycolipids. We show they are able to reproduce membrane properties and induce binding of peripheral membrane proteins. These test cases demonstrate the validity and transferability of our approach.

The carbohydrate glycosylphosphatidylinositol anchor chain under mechanical stress.

Carbohydrates have quite complicated micro heterogenic structure which may undergo different structural transitions. Due to their extreme flexibility it is very difficult to investigate such structural changes experimentally. In these studies we want to predict what structural and conformational changes are possible in the carbohydrate glycosylphosphatidylinositol anchor chain (GPI): the tetrasaccharide with the unique sequence Man-α(1 → 2)-Man-α(1 → 6)-Man-α(1 → 4)-GlcN-α. This is a very important biomolecule associated with the processes of transmitting various types of signals in the cells of living organisms. In order to investigate conformational and structural changes in GPI we use in these studies the theoretical Enforced Geometry Optimization (EGO) method. In this method a molecule is exposed to a mechanical stress caused by external forces applied to selected atoms. It turned out that under external stretching forces the mannopyranose unit can change its 4C1 chair conformation into three different forms: 1S3, oS2 and B2,5. The initial 4C1 glucosamine ring can transit into the twisted boat 1S3 and the boat Bo,3 conformations. The obtained results confirm the high flexibility of the GPI anchor sugar chain.

Silicon-bridged (1→1)-disaccharides: an umpoled glycomimetic scaffold.

Modification of the carbohydrate scaffold is an important theme in drug and vaccine discovery. Therefore, the preparation of novel types of glycomimetics is of interest in synthetic carbohydrate chemistry. In this manuscript, we present an early investigation of the synthesis, structure, and conformational behaviour of (1→1)-Si-disaccharides as a novel type of glycomimetics arising from the replacement of interglycosidic oxygen with a dimethyl-, methylpropyl-, or diisopropylsilyl linkage. We accomplished the preparation of this unusual group of umpoled compounds by the reaction of lithiated glycal or 2-oxyglycal units with dialkyldichlorosilanes. We demonstrated the good stability of the "Si-glycosidic" linkage under acidic conditions even at elevated temperatures. Next, we described the conformational landscape of these compounds by the combination of in silico modelling with spectroscopic and crystallographic methods. Finally, we explained the observed conformational flexibility of these compounds by the absence of gauche stabilizing effects that are typically at play in natural carbohydrates.

Structural characterization and antioxidant activity of Polygonatum sibiricum polysaccharides.

Polysaccharide is one of the main active components of Polygonatum sibiricum. For this study, P. sibiricum polysaccharides (PSP) were obtained through purification using DEAE-Cellulose52 and Sephacryl G-150 column chromatography. The obtained samples were named PSP1, PSP2 and PSP3. The PSP1 sample was found to have the highest content and the best solubility, and a subsequent. So, its structure and characterization were analyzed. The main sugar residue linkages were found to be â†’ 1)-ß-D-Fruf-(2 â†’ 1)-ß-D-Fruf-(2 â†’ 1), 1 â†’ -ß-D-Fruf-(2 â†’ 6)α-D-Glcp (1→, →4)-ß-D-Manp-(1 â†’ 4)-ß-D-Manp-(1→ and →6)-ß-D-Glcp-(1 â†’ 4)-ß-D-Manp-(1→ link existed. Branch chain analysis indicated →1,6)-ß-D-Fruf-(2→, ß-D-Fruf-(2→, →1,6)-ß-D-Fruf-(2→, →6)-ß-D- Fruf-(2→ link existed, and the link site was at position C-6. In vitro antioxidant activity tests showed that PSP1 had a certain scavenging effect on DPPH, hydroxyl radical, superoxide anion radical and a particular effect on the chelating ability of ferrous. This suggested that P. sibiricum polysaccharides may be a potential antioxidant.

N-Acetyl Side-Chain Conformation in Saccharides: Solution Models Obtained from MA'AT Analysis.

MA'AT analysis has been applied to model the conformational properties of N-acetyl side-chains in biologically important GlcNAc and ManNAc monosaccharides and in a ßGlcNAc-(1→4)-ßGlcNAc disaccharide. Density functional theory calculations were conducted to obtain parameterized equations that relate the magnitudes and signs of 10 spin-coupling constants to conformations of the C2-N2 bonds of GlcNAc and ManNAc. Six of these equations were used with experimental J-couplings, measured in H2O/2H2O and DMSO-d6 solvents in selectively 13C-labeled compounds, to model the C1-C2-N2-C1' torsion angle (θ1) in GlcNAc and ManNAc residues. MA'AT analysis gave mean values of θ1 of 106° for αGlcNAc and ∼116° for ßGlcNAc residues, with circular standard deviations (CSDs) of 21-22° in aqueous solution, in excellent agreement with those obtained by aqueous molecular dynamics (MD) simulation. Parameter space plots revealed unique MA'AT fits of the data, and root mean squared deviations (<0.2 Hz) were twofold smaller than those back-calculated from MD models, indicating that the MA'AT models better fit the experimental J-couplings. Context effects on both θ1 values were found to be small in a ßGlcNAc-(1→4)-ßGlcNAc disaccharide. MA'AT analysis gave a mean value of θ1 of 249° for αManNAc in H2O/2H2O, with a CSD of ∼19°, with both values in good agreement with MD. MA'AT models of N-acetyl side-chains are similar to those obtained previously for O-acetyl side-chains (J. Phys. Chem. B 2017, 121, 66-77). Both O- and N-acetylation conformationally constrain the C-O or C-N bonds relative to the same bonds in unsubstituted compounds. The present work confirms the ability of MA'AT analysis to reveal relatively small changes in mean molecular torsion angles in solution and provides additional evidence of the method as an experimental tool complementary to MD simulation.

Ferroptosis Related Immunomodulatory Effect of a Novel Extracellular Polysaccharides from Marine Fungus Aureobasidium melanogenum.

Marine fungi represent an important and sustainable resource, from which the search for novel biological substances for application in the pharmacy or food industry offers great potential. In our research, novel polysaccharide (AUM-1) was obtained from marine Aureobasidium melanogenum SCAU-266 were obtained and the molecular weight of AUM-1 was determined to be 8000 Da with 97.30% of glucose, 1.9% of mannose, and 0.08% galactose, owing to a potential backbone of α-D-Glcp-(1→2)-α-D-Manp-(1→4)-α-D-Glcp-(1→6)-(SO3-)-4-α-D-Glcp-(1→6)-1-ß-D-Glcp-1→2)-α-D-Glcp-(1→6)-ß-D-Glcp-1→6)-α-D-Glcp-1→4)-α-D-Glcp-6→1)-[α-D-Glcp-4]26→1)-α-D-Glcp and two side chains that consisted of α-D-Glcp-1 and α-D-Glcp-(1→6)-α-D-Glcp residues. The immunomodulatory effect of AUM-1 was identified. Then, the potential molecular mechanism by which AUM-1 may be connected to ferroptosis was indicated by metabonomics, and the expression of COX2, SLC7A11, GPX4, ACSL4, FTH1, and ROS were further verified. Thus, we first speculated that AUM-1 has a potential effect on the ferroptosis-related immunomodulatory property in RAW 264.7 cells by adjusting the expression of GPX4, regulated glutathione (oxidative), directly causing lipid peroxidation owing to the higher ROS level through the glutamate metabolism and TCA cycle. Thus, the ferroptosis related immunomodulatory effect of AUM-1 was obtained.

Practical non-enzymatic synthesis of propargyl sialyl-α-(2-3')-lactosamine trisaccharide using minimal protecting groups manipulation.

The trisaccharide, prop-2-ynyl 5-acetamido-3,5-dideoxy-d-glycero-α-d-galacto-2-nonulopyranosylonic acid-(2 â†’ 3)-ß-d-galactopyranosyl-(1 â†’ 4)-2-acetamido-2-deoxy-ß-d-glucopyranoside (9) has been efficiently synthesized in a few steps without the need of conformationally constrained glycosyl donors and acceptors or enzymes. First, using the known prop-2-ynyl 2-acetamido-2-deoxy-6-O-tert-butyldiphenylsilyl-ß-d-glucopyranoside as acceptor (2) and the peracetylated galactosyl trichloroacetimidate (3) as glycosyl donor, followed by protecting groups manipulation, prop-2-ynyl (6-O-tert-butyldiphenylsilyl-ß-d-galactopyranosyl)-(1 â†’ 4)-2-acetamido-2-deoxy-6-O-tert-butyldiphenylsilyl-ß-d-glucopyranoside (6) was synthesized with exclusive O-4 regioselectivity due to steric hindrance of the upper face of the acceptor at O-3. Sialylation with the thiophenyl glycosyl donor (7) afforded the desired trisaccharide with the shortest number of steps and in higher overall yield than previously reported methodologies. The direct use of minimally protected N-acetyl-lactosamine acceptor (6) was critical for the efficient synthesis of the title compound. The propargylic aglycone is suitable for chemical ligation using click chemistry as reported for its (2 â†’ 6) sialylated analog.

Carbohydrate Structure Database oligosaccharide conformation tool.

Population analysis in terms of glycosidic torsion angles is frequently used to reveal preferred conformers of glycans. However, due to high structural diversity and flexibility of carbohydrates, conformational characterization of complex glycans can be a challenging task. Herein, we present a conformation module of oligosaccharide fragments occurring in natural glycan structures developed on the platform of the Carbohydrate Structure Database. Currently, this module deposits free energy surface and conformer abundance maps plotted as a function of glycosidic torsions for 194 "inter"residue bonds. Data are automatically and continuously derived from explicit-solvent molecular dynamics (MD) simulations. The module was also supplemented with high-temperature MD data of saccharides (2,403 maps) provided by GlycoMapsDB (hosted by GLYCOSCIENCES.de project). Conformational data defined by up to 4 torsional degrees of freedom can be freely explored using a web interface of the module available at http://csdb.glycoscience.ru/database/core/search_conf.html.

The isolation, structural features and biological activities of polysaccharide from Ligusticum chuanxiong: A review.

Ligusticum chuanxiong, the dried rhizome of Ligusticum chuanxiong Hort, has been widely applied in traditional Chinese medicine for treating plague, and it has appeared frequently in the prescriptions against COVID-19 lately. Ligusticum chuanxiong polysaccharide (LCPs) is one of the effective substances, which has various activities, such as, anti-oxidation, promoting immunity, anti-tumor, and anti-bacteria. The purified fractions of LCPs are considered to be pectic polysaccharides, which are mainly composed of GalA, Gal, Ara and Rha, and are generally linked by α-1,4-d-GalpA, α-1,2-l-Rhap, α-1,5-l-Araf, ß-1,3-d-Galp and ß-1,4-d-Galp, etc. The pectic polysaccharide shows an anti-infective inflammatory activity, which is related to antiviral infection of Ligusticum chuanxiong. In this article, the isolation, purification, structural features, and biological activities of LCPs in recent years are reviewed, and the potential of LCPs against viral infection as well as questions that need future research are discussed.

Complete 1H and 13C NMR chemical shift assignments of mono-to tetrasaccharides as basis for NMR chemical shift predictions of oligo- and polysaccharides using the computer program CASPER.

Carbohydrate structure can be elucidated or confirmed by using NMR spectroscopy as the prime technique. Prediction of 1H and 13C NMR chemical shifts by computational approaches makes this assignment process more efficient and the program CASPER can perform this task rapidly. It does so by relying on chemical shift data of mono-, di-, and trisaccharides. In order to improve accuracy and quality of these predictions we have assigned 1H and 13C NMR chemical shifts of 30 monosaccharides, 17 disaccharides, 10 trisaccharides and one tetrasaccharide; in total 58 compounds. Due to different rotamers, ring forms, α- and ß-anomeric forms and pD conditions this resulted in 74 1H and 13C NMR chemical shift data sets, all of which were refined using total line-shape analysis for the 1H resonances in order to obtain accurate chemical shifts. Subsequent NMR chemical shift predictions for three sialic acid-containing oligosaccharides, viz., GD1a, a disialyl-LNnT hexasaccharide and a polysialic acid-lactose decasaccharide, and NMR-based structural elucidations of two O-antigen polysaccharides from E. coli O174 were performed by the CASPER program (http://www.casper.organ.su.se/casper/) resulting in very good to excellent agreement between experimental and predicted data thereby demonstrating its utility for carbohydrate compounds that have been chemically or enzymatically synthesized, structurally modified or isolated from nature.

SN- and NS-puckered sugar conformers are precursors of the (6-4) photoproduct in thymine dinucleotide.

Some amount of furanose in a southern conformation, possibly in both, but certainly in one of the two adjacent nucleotides of a dipyrimidine site, is necessary for (6-4) photoproduct formation in oligonucleotides. To explore the necessity, role, and most favorable location of each South sugar conformer in the formation of the (6-4) adduct in the thymine dinucleotide TpT, the photochemical behavior of two synthetic analogues, in which the South sugar conformation is prohibited for one of their two sugars, has been examined. Herein, we experimentally demonstrate that the presence of one sugar presenting some amount of South puckering, at any of the extremities, is sufficient to trigger (6-4) adduct formation. Nonetheless, the photochemical behavior of the dinucleotide with a South-puckered conformation at the 5'-end, mimics more closely that of TpT. In addition, using the 5' North 3' South-dilocked dinucleotide, we demonstrate that the flexibility of the South pucker at the 3'-end has little influence on the (6-4) adduct formation.