Using the P4 unknotting assay, DNA topoisomerase I1 has been purified from several mammalian cells. Similar to prokaryotic DNA gyrase, mammalian DNA topoisomerase I1 can cleave double-stranded DNA and be trapped as a covalent protein-DNA complex. This cleavage reaction requires protein denaturant treatment of the topoisomerse 11-DNA complex and is reversible with respect to salt and temperature...

FokI is a type IIs restriction endonuclease comprised of a DNA recognition domain and a catalytic domain. The structural similarity of the FokI catalytic domain to the type II restriction endonuclease BamHI monomer suggested that the FokI catalytic domains may dimerize. In addition, the FokI structure, presented in an accompanying paper in this issue of Proceedings, reveals a dimerization inter...

BACKGROUND Inhaled rhDNase may improve sputum viscosity and mucociliary clearance by cleavage of extracellular DNA derived for instance from dead leukocytes in purulent, highly viscous patient sputum. METHODS Here we established a method to quantify rhDNase-mediated DNA fragmentation in sputum using gel electrophoresis. Sputum of Pseudomonas aeruginosa colonized cystic fibrosis (CF) patients ...

During cut-and-paste mariner/Tc1 transposition, transposon DNA is cut precisely at its junction with flanking DNA, ensuring the transposon is neither shortened nor lengthened with each transposition event. Each transposon end is flanked by a TpA dinucleotide: the signature target site duplication of mariner/Tc1 transposition. To establish the role of this sequence in accurate DNA cleavage, we h...

Topoisomerase II resolves intrinsic topological problems of double-stranded DNA. As part of its essential cellular functions, the enzyme generates DNA breaks, but the regulation of this potentially dangerous process is not well understood. Here we report single-molecule fluorescence experiments that reveal a previously uncharacterized sequence of events during DNA cleavage by topoisomerase II: ...

Microcin B17 is a 3.1-kDa bactericidal peptide; the putative target of this antibiotic is DNA gyrase. Microcin B17 has no detectable effect on gyrase-catalysed DNA supercoiling or relaxation activities in vitro and is unable to stabilise DNA cleavage in the absence of nucleotides. However, in the presence of ATP, or the non-hydrolysable analogue 5'-adenylyl beta,gamma-imidodiphosphate, microcin...

Following V(D)J cleavage, the newly liberated DNA signal ends can be either fused together into a signal joint or used as donor DNA in RAG-mediated transposition. We find that both V(D)J cleavage and release of flanking coding DNA occur before the target capture step of transposition can proceed; no coding DNA is ever detected in the target capture complex. Separately from its role in V(D)J cle...

Abstract Thermophilic Argonaute proteins (Agos) have been shown to utilize small DNA guides for cleaving complementary in vitro, which shows great potential nucleic acid detection. In this study, we explored mesophilic Agos the detection of molecule by cooperating with allosteric transcription factors (aTFs). Two from bacteria, Paenibacillus borealis ( Pb Ago) and Brevibacillus laterosporus Bl ...

Cytotoxic T cells (CTL) represent the major defense mechanism against the spread of virus infection. It is believed that the pore-forming protein, perforin, facilitates the entry of a series of serine proteases (particularly granzyme B) into the target cell which ultimately leads to DNA fragmentation and apoptosis. We demonstrate here that during CTL-mediated cytolysis the catalytic subunit of ...

Two acridine groups were successfully introduced into di-iron(III) complex. DNA cleavage experiments indicated that complex conjugating bisacridine groups can enhance 300-fold for the cleavage efficiency compared with complex lacking of acridine conjugation. Further ligation assay of DNA segments provided the evidence for hydrolytic mechanism of DNA cleavage.