ABSTRACT
Protein synthesis, which takes place within ribosomes, is essential for the survival of any living organism. Ribosomes are composed of both proteins and RNA. Specific interaction between the 3' end CCUCC sequence of prokaryotic 16S rRNA and a partially complementary sequence preceding the initiating codon of mRNA is believed to be a prerequisite for initiation of protein synthesis. Here we report the use of short (three to six nucleotides) synthetic DNA analogs complementary to this sequence to block protein synthesis in vitro and in vivo in Escherichia coli. In the DNA analogs the normal phosphodiester bond in the antisense DNA was replaced by methylcarbamate internucleoside linkages to enhance transport across plasma membranes. Of the analogs tested, those with the sequence AGG and GGA inhibit protein synthesis and colony formation by E. coli strains lacking an outer cell wall. Polyethylene glycol 1000 (PEG 1000) was attached to the 5' end of some of the test methylcarbamate DNAs to enhance solubility. Analogs of AGG and GGAG with PEG 1000 attached inhibited colony formation in normal E. coli. These analogs may be useful food additives to control bacterial spoilage and biomedically as antibiotics.
Subject(s)
Anti-Bacterial Agents/pharmacology , DNA, Antisense/pharmacology , Escherichia coli/drug effects , Oligonucleotides, Antisense/pharmacology , Protein Synthesis Inhibitors/pharmacology , RNA, Messenger/genetics , RNA, Ribosomal, 16S/genetics , 3T3 Cells , Animals , Base Sequence , Cell Division/drug effects , Escherichia coli/genetics , Escherichia coli/metabolism , Mice , Molecular Sequence Data , Protein Biosynthesis/drug effectsABSTRACT
A novel oligonucleotide analog has been prepared from ribonucleoside derived morpholine subunits linked by carbamate groups. Oxidative cleavage of the 2',3' vicinal diol of cytidine followed by reductive amination of the resulting dialdehyde afforded the morpholine subunit. Coupling of the subunits are through carbamate moieties and the oligomers were characterized by 1H NMR and FAB MS. Evidence for interaction of the hexamer 19 with p(dG6) was found, but an atypical interaction of 19 with a RNA target was observed.
Subject(s)
Carbamates , Morpholines/chemical synthesis , Oligonucleotides/chemical synthesis , Chemical Phenomena , Chemistry , Chromatography , Hot Temperature , Hydrogen-Ion Concentration , Magnetic Resonance Spectroscopy , Mass Spectrometry , Molecular Structure , Nucleic Acid Denaturation , SolubilityABSTRACT
Oligonucleotide analogs, in which carbamate rather than phosphodiester linkages form the backbone, have been analyzed by negative ion fast atom bombardment mass spectrometry. These oligodeoxynucleotides, dimers (982.4 daltons) to 11-mers (4356.5 daltons), show intense [M - H]- ions and some degree of cleavage of the sugar-carbamate backbone structure in repeating fashion on both sides of the carbonyl groups. Sequencing of the shorter chain oligonucleotide analogs, up to six bases long, is demonstrated by complete 3' and 5' terminal sequence fragment ions. Longer chain oligomers show partial sequencing information.
