ABSTRACT
Conjugate DNAzymes with polyamines and peptides were successfully prepared by solid phase fragment condensation (SPFC) and showed up to 4.2 times higher catalytic efficiency (k(cat)/K(m)) and enhanced tolerance against DNase 1digestion. To be pointed out, intracellular localization of DNAzymes could be controlled by conjugated with naturally occurring signal peptides responsible for nuclear cytoplasmic transport of proteins.
Subject(s)
DNA, Catalytic/metabolism , RNA, Viral/metabolism , Base Sequence , Gene Products, gag/genetics , HIV-1/genetics , Hydrolysis , RNA, Viral/chemistryABSTRACT
Precisely controlled intracellular delivery of oligonucleotides was achieved by conjugation with signal peptides and amphiphilic designed peptides. Localization in the cellular nucleus was accelerated by nuclear localizing signals derived from naturally occurring viral proteins and arginine rich designed peptides, and cytoplasmic localization was enhanced by nuclear export signals and membrane fusion peptides.
Subject(s)
DNA/metabolism , Drug Delivery Systems , Peptides/metabolism , Amino Acid Sequence , Cell Death/drug effects , DNA/chemical synthesis , Deoxyribonucleases/metabolism , Humans , Jurkat Cells , Molecular Sequence Data , Oligonucleotides, Antisense/pharmacology , Peptides/chemical synthesis , Peptides/chemistry , Ribonuclease H/metabolism , Telomerase/metabolismABSTRACT
In the present study, membrane permeability and intracellular localization of oligonucleotide (ODN) conjugated with naturally occurring functional peptides or designed peptide were investigated, as well as antisense properties of them to inhibit of telomerase activities. All conjugate antisense ODNs showed higher membrane permeability and nuclease resistance than natural ODN. Intracellular localization of ODN could be precisely controlled by conjugation with functional peptides. Conjugate antisense ODNs indicated thousand fold higher inhibitory effects than natural ODN in cellular extract and 95% suppression in human leukemia cells.
Subject(s)
Gene Expression , Gene Transfer Techniques , Oligonucleotides, Antisense/metabolism , Protein Sorting Signals , Base Sequence , Cell Extracts , Cell Survival/drug effects , Gene Expression/drug effects , Humans , Jurkat Cells , Molecular Sequence Data , Oligonucleotides, Antisense/genetics , Oligonucleotides, Antisense/pharmacology , Telomerase/antagonists & inhibitorsABSTRACT
Glycosidase-promoted hydrolysis was performed in a microreaction channel. The result was compared with the reaction in a micro-test tube. Transgalactosylation on p-nitrophenyl-2-acetamide-2-deoxy-beta-D-glucopyranoside was also examined in a microreaction channel, because transglycosylation is a useful method for oligosaccharide synthesis. We examined both the forward reaction, i.e., hydrolysis, and the reverse reaction, i.e., transglycosylation, in the microreaction channel. The results showed that both hydrolysis and transglycosylation were enhanced in the microreaction channel compared with the batch reaction.
