Restriction of intracellular replication by restoring TFEB-mediated xenophagy.

Unlabelled: Macroautophagy/autophagy functions as a part of the innate immune system in clearing intracellular pathogens. Although this process is well known, the mechanisms that control antibacterial autophagy are not clear. In this study we show that during intracellular infection, the activity of TFEB (transcription factor EB), a master regulator of autophagy and lysosome biogenesis, is suppressed by maintaining it in a phosphorylated state on the lysosomes.

Ca(2+)-mediated site-specific DNA cleavage and suppression of promiscuous activity of KpnI restriction endonuclease.

The characteristic feature of type II restriction endonucleases (REases) is their exquisite sequence specificity and obligate Mg(2+) requirement for catalysis. Efficient cleavage of DNA only in the presence of Ca(2+) ions, comparable with that of Mg(2+), is previously not described. Most intriguingly, KpnI REase exhibits Ca(2+)-dependent specific DNA cleavage.

KpnI restriction endonuclease and methyltransferase exhibit contrasting mode of sequence recognition.

The molecular basis of the interaction of KpnI restriction endonuclease (REase) and the corresponding methyltransferase (MTase) at their cognate recognition sequence is investigated using a range of footprinting techniques. DNase I protection analysis with the REase reveals the protection of a 14-18 bp region encompassing the hexanucleotide recognition sequence. The MTase, in contrast, protects a larger region.

Kinetic and catalytic properties of dimeric KpnI DNA methyltransferase.

KpnI DNA-(N(6)-adenine)-methyltransferase (KpnI MTase) is a member of a restriction-modification (R-M) system in Klebsiella pneumoniae and recognizes the sequence 5'-GGTACC-3'. It modifies the recognition sequence by transferring the methyl group from S-adenosyl-l-methionine (AdoMet) to the N(6) position of adenine residue. KpnI MTase occurs as a dimer in solution as shown by gel filtration and chemical cross-linking analysis.