The recognition of 'regular' and 'oxidized' sites of base loss (AP sites) in DNA by various AP endonucleases was compared. Model substrates with regular AP sites (resulting from mere hydrolysis of the glycosylic bond) were produced by damaging bacteriophage PM2 DNA by exposure to low pH; those with AP sites oxidized at the C-4'- and C-1'-position of the sugar moiety by exposure to Fe(III)-bleom...

The RTH1 nuclease is involved in the replication of chromosomal DNA as well as in the repair of DNA damage. Replication protein A (RPA) is also an integral part of the DNA replication and repair processes. We have investigated the roles(s) of RPA in the function of RTH1 nuclease, including its structure specific endonuclease activity. Initial in vitro studies, which employed a "flap" or a "pseu...

The mitochondrial genome is highly susceptible to damage by reactive oxygen species (ROS) generated endogenously as a byproduct of respiration. ROS-induced DNA lesions, including oxidized bases, abasic (AP) sites, and oxidized AP sites, cause DNA strand breaks and are repaired via the base excision repair (BER) pathway in both the nucleus and mitochondria. Repair of damaged bases and AP sites i...

Homing endonucleases are enzymes that catalyze the highly sequence-specific cleavage of DNA. We have developed an in vivo selection in Escherichia coli that links cell survival with homing endonuclease-mediated DNA cleavage activity and sequence specificity. Using this selection, wild-type and mutant variants of three homing endonucleases were characterized without requiring protein purificatio...

We have discovered a new restriction endonuclease, MfeI, in nuclear extracts from T cells contaminated with Mycoplasma fermentans. This endonuclease was identified while studying proteins binding to the interleukin-2 receptor alpha chain gene promoter. MfeI cuts at the recognition sequence C'AATTG generating EcoRI compatible cohesive ends. Potential applications are discussed.

We have developed a highly sensitive and selective fluorescence polarization assay method based on the specificity of the DNA cleavage reaction with the enhancement of gold nanoparticles (AuNPs) for assaying endonuclease activity and inhibition. This assay can detect EcoRI endonuclease down to 5.0×10(-4) U mL(-1) with a detection range from 5.0×10(-4) to 10 U mL(-1).

A novel sequence-specific endonuclease has been isolated from Streptococcus cremoris F. ScrFI recognises the sequence: (formula; see text) and cleaves as indicated by the arrow ( ). It is the first enzyme to recognise this sequence and the first endonuclease reported from the lactic streptococci used in dairy fermentations.

MmeI is an unusual Type II restriction enzyme that is useful for generating long sequence tags. We have cloned the MmeI restriction-modification (R-M) system and found it to consist of a single protein having both endonuclease and DNA methyltransferase activities. The protein comprises an amino-terminal endonuclease domain, a central DNA methyltransferase domain and C-terminal DNA recognition d...

An important biochemical hallmark of apoptosis is the cleavage of chromatin into oligonucleosomal fragments. Here, we purified a Mg2+-dependent endonuclease from etoposide-treated HL-60 cells undergoing apoptosis. High levels of Mg2+-dependent endonuclease activity were detected in etoposide-treated HL-60 cells, and this activity increased in a time-dependent manner following etoposide treatmen...