Biology subjects

Fatma, S.

Publications and source records attributed to Fatma, S..

3 recordsLinked to original sources

bioRxiv2021

Sequential rescue and repair of stalled and damaged ribosome by bacterial PrfH and RtcB

In bacteria, rescue of stalled ribosomes due to 3-truncated mRNAs is carried out by the ubiquitous trans-translation system as well as alternative ribosome-rescue factors such as ArfA and ArfB. It is unclear, however, how the stalled ribosomes caused by ribosomal damages are rescued. Here, we report that a bacterial system composed of PrfH and RtcB not only rescues a stalled ribosome resulting from a specific damage in the decoding center but also repairs the damage afterwards. Peptide release assays reveal that PrfH is only active with the damaged ribosome, but not with the intact one. A 2.55-angstrom cryo-EM structure of PrfH in complex with the damaged 70S ribosome provides molecular insight into specific recognition of the damage site by PrfH. RNA repair assays demonstrate that PrfH-coupled RtcB efficiently repairs the damaged 30S ribosomal subunit, but not the damaged tRNAs. Thus, our studies have uncovered a biological operation by a pair of bacterial enzymes, aiming to reverse the potentially lethal damage inflicted by an invading ribotoxin for cell survival.

biochemistry↗

bioRxiv2021

Molecular mechanisms of the CdnG-Cap5 antiphage defense system employing 3',2'-cGAMP as the second messenger

Cyclic-oligonucleotide-based antiphage signaling systems (CBASS) are diverse and abundant in bacteria. Here, we present biochemical and structural characterization of two CBASS systems, composed of CdnG and Cap5, from Asticcacaulis sp. and Lactococcus lactis. We show that CdnG from Asticcacaulis sp. synthesizes 3,2-cGAMP in vitro, and 3,2-cGAMP is the biological signaling molecule that activates Cap5 for DNA degradation. Crystal structures of Cap5, together with the SAVED domain in complex with 3,2-cGAMP, provide insight into the architecture of Cap5 as well as molecular recognition of 3,2-cGAMP by the SAVED domain of Cap5. Amino acid conservation of the SAVED domain of Cap5, together with mutational studies, led us to propose a novel mechanism of Back-to-Front stacking of two SAVED domains, mediated by 3,2-cGAMP, to activate HNH nuclease domain for DNA degradation. Our study of the most abundant CBASS system provides new insight into mechanisms employed by bacteria in their conflicts against phage.

biochemistry↗

bioRxiv2020

epiGBS2: an improved protocol and automated snakemake workflow for highly multiplexed reduced representation bisulfite sequencing

epiGBS is an existing reduced representation bisulfite sequencing method to determine cytosine methylation and genetic polymorphisms de novo. Here, we present epiGBS2, an improved epiGBS laboratory protocol and user-friendly bioinformatics pipeline for a wide range of species with or without reference genome. epiGBS2 decreases costs and time investment and increases user-friendliness and reproducibility. The library protocol was adjusted to allow for a flexible choice of restriction enzymes and a double digest. Instead of fully methylated adapters, semi-methylated adapters are now used. The bioinformatics pipeline was improved in speed and integrated in the snakemake workflow management system, which now makes the pipeline easy to execute, modular, and parameter settings flexible. We also provide a detailed description of the laboratory protocol, an extensive manual of the bioinformatics pipeline, which is publicly accessible on github (https://github.com/nioo-knaw/epiGBS2) and zenodo (https://doi.org/10.5281/zenodo.3819996), and example output.

bioinformatics↗