A Convenient Oligonucleotide Conjugation via Tandem Staudinger Reaction and Amide Bond Formation at the Internucleotidic Phosphate Position.

Staudinger reaction on the solid phase between an electronodeficit organic azide, such as sulfonyl azide, and the phosphite triester formed upon phosphoramidite coupling is a convenient method for the chemical modification of oligonucleotides at the internucleotidic phosphate position. In this work, 4-carboxybenzenesulfonyl azide, either with a free carboxy group or in the form of an activated ester such as pentafluorophenyl, 4-nitrophenyl, or pentafluorobenzyl, was used to introduce a carboxylic acid function to the terminal or internal internucleotidic phosphate of an oligonucleotide via the Staudinger reaction. A subsequent treatment with excess primary alkyl amine followed by the usual work-up, after prior activation with a suitable peptide coupling agent such as a uronium salt/1-hydroxybenzotriazole in the case of a free carboxyl, afforded amide-linked oligonucleotide conjugates in good yields including multiple conjugations of up to the exhaustive modification at each phosphate position for a weakly activated pentafluorobenzyl ester, whereas more strongly activated and, thus, more reactive aryl esters provided only single conjugations at the 5'-end. The conjugates synthesized include those with di- and polyamines that introduce a positively charged side chain to potentially assist the intracellular delivery of the oligonucleotide.

Phase transitions in the crystals of L- and DL-cysteine on cooling: the role of the hydrogen-bond distortions and the side-chain motions. 2. DL-cysteine.

Structural strain and a first-order phase transition in the crystalline DL-cysteine on cooling and on reverse heating were followed by Raman spectroscopy and X-ray diffraction. The transition is reversible and has a large hysteresis (over 100 K). The temperature at which the transition is observed depends strongly on the cooling/heating rate. The phase transition is accompanied by crystal fragmentation. The low-temperature phase could be obtained not only as a result of the solid-state transformation in situ as a polycrystalline sample (with strong preferred orientation, or without it, depending on the preparative technique), but also (using an original crystallization technique) as a single crystal of the quality suitable for structural analysis. For the first time, the crystal structure of the low-temperature phase was solved independently by powder and by single-crystal diffraction techniques. The spectral changes were correlated with the precise diffraction data on the intramolecular conformations and the intermolecular hydrogen bonding before and after the phase transition. The role of the distortion of the intermolecular hydrogen bonds and of the motions of the -CH(2)SH side chains in the phase transition is discussed in a comparison with the low-temperature phase transition in L-cysteine, which is of a different type and preserves the single crystals intact (Kolesov et al. J. Phys. Chem. B, 2008, 112 (40), 12827-12839).

Pressure-induced phase transitions in crystalline L- and DL-cysteine.

A series of extended reversible phase transitions at approximately 0.1, 1.5, 2.0, and approximately 5 GPa was observed for the first time in the crystals of dl-cysteine by Raman spectroscopy. These are the first examples of the phase transitions induced by increasing pressure in the racemic crystal of an amino acid. In the crystals of the orthorhombic l-cysteine, a sequence of reversible structural changes in the pressure range between 1.1 and 3 GPa could be observed by Raman spectroscopy, instead of a single sharp phase transition at 1.9 GPa reported previously in ( Moggach, et al. Acta Crystallogr. 2006, B62, 296- 309 ). The role of the movements of the side -CH 2SH groups and of the changes in the hydrogen-bonding type in dl- and l-cysteine during the phase transitions with increasing pressure is discussed and compared with that on cooling down to 3 K.