Ashwani K. Sharma

Conformations of peptides containing Z-alpha,beta-dehydroleucine (delta ZLeu). A comparison of Boc-Pro-delta ZLeu-Gly-NHEt and Boc-Pro-delta ZPhe-Gly-NHEt.

Two tripeptides of the type Boc-Pro-delta ZX-Gly-NHEt (where X = Leu, Phe) have been synthesized and their solution conformations investigated by 270 MHz 1H n.m.r. and i.r. spectroscopy. These conformational studies indicated that delta ZLeu, similar to delta ZPhe, has a strong tendency to stabilize folded Type II beta-turn conformations when present at i + 2 position.

Microwave-assisted spectrophotometric estimation of polymer-supported functional groups using a universal reagent

A rapid method has been developed for the estimation of polymer-supported functionalities under microwave irradiation. The method involves the use of a novel universal reagent, S-(4,4′-dimethoxytrityl)-3-mercaptopropionic acid (DMPA) for the estimation of polymer-supported hydroxyalkyl, aminoalkyl and mercaptoalkyl functionalities in the presence of triphenylphosphine–bromotrichloromethane (TPP–BTCM) as an oxidation-reduction coupling reagent. The loadings obtained on the supports following the proposed method were found to be comparable with those obtained with the standard, 4,4′-dimethoxytrityl chloride (DMTr-Cl), method. The usefulness of the method was further demonstrated by monitoring the functionalization of polymer supports, suitable for solid-phase peptide and oligonucleotide synthesis.

Composite Magnetic and Non-magnetic Beads as Efficient Solid Supports for Machine-Aided Oligonucleotide Synthesis

by Pradeep Kumar, Ashwani K. Sharma, and Kailash C. Gupta*Nucleic Acids Research Laboratory, Centre for Biochemical Technology, Mall Road, Delhi University Campus,Delhi-110007, Indiaand Shlomo Margel and Sigalit GuraDepartment of Chemistry, Bar-Ilan University, Ramat-Gan 52900, Israeland Hartmut SeligerUniversitat Ulm, Sektion Polymere, Albert Einstein Allee 11, D-89069 UlmThe synthesis of oligodeoxyribonucleotides on composite magnetic and nonmagnetic beads of 4.5 mm witha cleavable and non-cleaveable linker for biochemical applications is described.

Preparation of high-loading polymer supports by polymerization reaction useful for oligonucleotide synthesis

A simple protocol based on polymerization reactions has been developed for the preparation of high-loading polymer supports, useful for large-scale synthesis of oligonucleotides. Polymer supports of different pore sizes have been employed in the present investigation to improve the functional-group density on them. A ten- to twelvefold increase in the loading of the functional groups, after the polymerization reaction, has been observed. The support was then used in the subsequent reaction to attach the leader nucleoside to obtain fully functionalized supports 6a–6c by oligonucleotide synthesis in an automated DNA synthesizer. The aminoalkylated-supports 5a–5c were directly employed for the synthesis of oligonucleotide 3′-phosphates. The oligonucleotides and oligonucleotide 3′-phosphates synthesized on these supports were compared with the corresponding standard oligomers with respect to their retention time on HPLC. These were further characterized on MALDI-TOF mass spectrometry.

High-Loading Supports for Oligonucleotide Synthesis

b ), and A simple protocol for the preparation of high-loading supports, suitable for large-scale synthesis of oligonucleotides, has been developed. The method involves the use of inexpensive reagents and is amenable to large-scale preparation of supports. The derivatized supports were successfully employed in an automated DNA synthesizer without any difficulty. The quality of the synthesized oligonucleotides was found to be comparable to that of the corresponding oligomers prepared with commercially available standard supports. Introduction. - Synthetic oligonucleotides and their modified analogs have become vital tools for various biological studies. They are finding widespread use in the development of oligonucleotide-based therapeutic and diagnostic tools (1 - 4). Gen- erally, oligonucleotides, in the size range of 15 - 30 residues, are required in abundance, but in tiny quantities for studies like DNA sequencing, etc. However, in case of developing DNA based diagnostics and therapeutics (for use as antisense oligomers), relatively larger quantities of short oligomers are required. To synthesize these molecules in large quantities, the present day methodologies (5 - 7) have proven to be somewhat expensive. Hence, there is a need to develop economically viable routes or protocols for the synthesis of oligonucleotides to meet this exponentially growing demand for these molecules. In the last several years, tremendous improvements have been reported in the area of oligonucleotide synthesis (8) (9). The introduction of new sets of labile protecting groups, fast coupling reagents, and improved polymer supports have revolutionized the entire field so that even a non-chemist can synthesize these molecules quite easily. Since most of the syntheses are currently carried out on solid supports, which play a significant role in deciding the cost of these molecules, there is a need to develop supports having a relatively higher density of the functional groups, which could ultimately reduce the cost of these molecules without compromising the quality of the final product. During the last few years, however, although several polymer supports have been developed and tested, LCAA-CPG (long-chain alkyl- amine/controlled-pore glass) is still the standard polymer support (10) for routine synthesis of oligonucleotides. Recently, we published an express protocol for the functionalization of polymer supports useful for oligonucleotide synthesis (12), in which we reported the loading of LCAA-CPG in the range of 45 - 50 mmol/g of support.