Synthesis and triplex-forming properties of cyclic oligonucleotides with (G,A)-antiparallel strands

Cyclic oligonucleotides carrying an oligopurine Watson-Crick sequence linked to the corresponding (G,A)- and (G,T)-antiparallel strands were prepared by nonenzymatic template-assisted cyclization of phosphorylated precursors. Cyclization was attempted using 3'-phosphate and 5'-phosphate linear precursors with carbodiimide or BrCN activation. The best results were obtained with the 5'-phosphorylated precursors and carbodiimide activation. Cyclic oligonucleotides bind polypyrimidine target sequence by formation of antiparallel triplexes. We have used UV and circular dichroism (CD) spectroscopy to analyze triplexes formed by cyclic oligonucleotides carrying G and A in the reverse-Hoogsteen strand. The relative stability of the triplexes formed by cyclic and linear oligonucleotides with a common polypyrimidine target was determined by melting experiments. The most-stable triplexes were formed by the cyclic oligonucleotide, followed by the unphosphorylated and phosphorylated oligonucleotide precursors, and, finally, the corresponding hairpin. Although the differences in binding affinity between cyclic oligonucleotides and their corresponding linear precursors are small, the use of cyclic oligonucleotides offers a clear advantage over conventional duplex recognition.     

Role of lipid protrusions in the function of interfacial enzymes

Secretory phospholipase A₂ (sPLA₂) is a class of interfacially active enzymes that selectively hydrolyze lipid molecules organized at interfaces like membranes. We present a simple theoretical model that relates the sPLA₂ action to the protrusions of the lipid molecules. The model explains (1) the observed enhancement of enzymatic activity by lipids with flexible, neutral, water-soluble polymers linked to their head groups and (2) the lag-burst kinetics of sPLA₂. It yields qualitative predictions of the effect of the initial composition of the membrane, the molecular weight of the polymer, and the composition of the hydrolysis products.     

Synthesis of oligonucleotide-peptide conjugates carrying the c-myc peptide epitope as recognition system

Oligonucleotide-peptide conjugates 1-3 were prepared by sequential addition of the appropriate Boc-protected amino acids, followed by nucleoside phosphoramidites in the same support. These molecules are designed to be used for triplex formation and for the directed assembly of nanomaterials. The structures of the desired oligonucleotide-peptide conjugates were confirmed by mass spectrometry on small oligonucleotide-peptide conjugates, by gel electrophoresis, and by hybridization with complementary oligonucleotides. Oligonucleotides carrying the c-myc peptide were specifically recognized by the anti-c-myc monoclonal antibody.     

Casein-derived phosphopeptides disrupt tight junction integrity,and precipitously dry up milk secretion in goats

Mammary involution is triggered by local stimuli, but the precise mechanism has not been defined. Milk stasis accumulate local signals, which makes the tight junctions (TJ) leaky. The aim of the study was to check the hypothesis that casein hydrolyzates (CNH) compromise TJ integrity and dry up milk secretion. A single dose of CNH transiently (12 to 24 h) compromised TJ integrity in the treatedudder. This was associated by a transient (12 to 96 h) decline in milk secretion. No such changes were recorded in the contralateral gland that served as a control. Four repeated doses of CNH after each milking caused drastic changes in mammary secretion and composition, which were associated with irreversible cessation of milk secretion within 96 h. No such changes were recorded in goats treated with de-phosphorylated casein (control). We conclude that CNH are the milk-borne factors that cause the disruption of TJ integrity and induction of involution, and that the serine-Ps in the CNHs are essential for the excretion of biological activity.     

Triplex Formation Using Oligonucleotide Clamps Carrying 8-Aminopurines

The synthesis and properties of triplex-forming DNA clamps carrying 8-aminopurines are described. The stability of triple helices is enhanced by replacing purine bases with 8-aminopurine residues. These enhanced binding properties are used for the specific capture of polypyrimidine RNA/DNA sequences of interest.