Histidine-Triad Hydrolases Provide Resistance to Peptide-Nucleotide Antibiotics.

The Escherichia coli microcin C (McC) and related compounds are potent Trojan horse peptide-nucleotide antibiotics. The peptide part facilitates transport into sensitive cells. Inside the cell, the peptide part is degraded by nonspecific peptidases releasing an aspartamide-adenylate containing a phosphoramide bond. This nonhydrolyzable compound inhibits aspartyl-tRNA synthetase. In addition to the efficient export of McC outside the producing cells, special mechanisms have evolved to avoid self-toxicity caused by the degradation of the peptide part inside the producers. Here, we report that histidine-triad (HIT) hydrolases encoded in biosynthetic clusters of some McC homologs or by standalone genes confer resistance to McC-like compounds by hydrolyzing the phosphoramide bond in toxic aspartamide-adenosine, rendering them inactive. IMPORTANCE Uncovering the mechanisms of resistance is a required step for countering the looming antibiotic resistance crisis. In this communication, we show how universally conserved histidine-triad hydrolases provide resistance to microcin C, a potent inhibitor of bacterial protein synthesis

DESCRIPTION OF A NEW POTENTIAL AGGREGATION FACTOR FROM THE STREPTOCOCCUS THERMOPHILUS GENOME

Autoaggregation, the ability to self-aggregate, is widespread among both Gram-positive and Gram-negative bacteria. The functional role of aggregation is not fully understood, but it is believed to be involved in the adaptation of bacteria to environmental conditions (PMID: 31294207). One interesting class of compounds responsible for the aggregation of lactic acid bacteria is aggregation factors—surface high-molecular-weight proteins rich in threonine and lysine (PMID: 30027759). Recently, our research group discovered a new strain of Streptococcus thermophilus in the mountainous regions of Serbia, exhibiting an aggregation phenotype. Aggregation phenotype was confirmed visually and using microscopy. Complete genome of Agg+ strain was sequenced using NGS and a gene encoding a potential aggregation factor, which was named aggS was identified. The predicted threonine (12.5%) and lysine (10.5%) rich protein contains 2367 amino acids, with an average molecular weight of 255986.63 Da. AggS also contains two cysteine residues,whereas previously well-described aggregation factors of this type did not contain any cysteine residues. The predicted protein includes an N-terminal YSIRK-like signal sequence and an LPXTG cell wall anchor domain. It has 6 Mucin binding domain repeats alternating with 6 Mub B2-like domain repeats. Additionally, we found a region resembling an ice-binding domain. Given that these bacteria endure prolonged periods of low temperatures, it can be speculated that this surface membrane protein also helps the bacteria withstand freezing. The fact that the alignment using BLASTp revealed AggS to be most closely related to an uncharacterised protein from the genome of Lactococcus garvieae, along with the discovery of a transposase gene sequence upstream of the gene, suggests that the aggregation factor was likely acquired through horizontal gene transfer. We plan to clone it into a shuttle vector and investigate the aggregation phenotype using a heterologous expression system in Lactococcus lactis, as well as explore its other functions.Book of abstract: From biotechnology to human and planetary health XIII congress of microbiologists of Serbia with international participation Mikromed regio 5, ums series 24: 4th – 6th april 2024, Mona Plaza hotel, Belgrade, Serbi

СЛУЧАЙ МИОПАТИИ ЭРБА-РОТА

The article presents a clinical case of a patient with Erb-Roth’s hereditary neuromuscular dystrophy presented with severe cardiomyopathy.В статье представлен клинический случай развития тяжелой формы кардиомиопатии у пациента, страдающего наследственным нейромышечным заболеванием – миопатией Эрба-Рота

A Novel Laser-Based Zebrafish Model for Studying Traumatic Brain Injury and Its Molecular Targets

Traumatic brain injury (TBI) is a major public health problem. Here, we developed a novel model of non-invasive TBI induced by laser irradiation in the telencephalon of adult zebrafish (Danio rerio) and assessed their behavior and neuromorphology to validate the model and evaluate potential targets for neuroreparative treatment. Overall, TBI induced hypolocomotion and anxiety-like behavior in the novel tank test, strikingly recapitulating responses in mammalian TBI models, hence supporting the face validity of our model. NeuN-positive cell staining was markedly reduced one day, but not seven days, after TBI, suggesting increased neuronal damage immediately after the injury, and its fast recovery. The brain-derived neurotrophic factor (Bdnf) level in the brain dropped immediately after the trauma, but fully recovered seven days later. A marker of microglial activation, Iba1, was elevated in the TBI brain, albeit decreasing from Day 3. The levels of hypoxia-inducible factor 1-alpha (Hif1a) increased 30 min after the injury, and recovered by Day 7, further supporting the construct validity of the model. Collectively, these findings suggest that our model of laser-induced brain injury in zebrafish reproduces mild TBI and can be a useful tool for TBI research and preclinical neuroprotective drug screening. © 2022 by the authors.Saint Petersburg State University, SPbU: 73026081; Russian Science Foundation, RSF: 20-65-46006We thank Alisa S. Belova for technical support in experimental manipulations and cortisol assay. We also thank Anatoly A. Maslov for the idea of using laser radiation to introduce brain damage. A.V.K. lab is supported by St. Peterburg State University funds (Pure ID 73026081).This study was funded by Russian Science Foundation (grant No. 20-65-46006)

The Relationship between the Strength Characteristics of Cerebral Aneurysm Walls with Their Status and Laser-Induced Fluorescence Data

Background: Cerebral aneurysms (CA) are a widespread vascular disease affecting 50 per 1000 population. The study of the influence of histological, morphological and hemodynamic factors on the status of the aneurysm has been the subject of many works. However, an accurate and generally accepted relationship has not yet been identified. Methods: In our work, the results of mechanical and spectroscopic measurements are considered. Total investigated 14 patients and 36 their samples of CA tissue. Results: The excitation–emission matrix of each specimen was evaluated, after which the strength characteristics of the samples were investigated. Conclusions: It has been shown that there is a statistically significant difference in the size of the peaks of two components, which characterizes the status of the aneurysms. In addition, a linear regression model has been built that describes the correlation of the magnitude of the ultimate strain and stress with the magnitude of the peaks of one of the components. The results of this study will serve as a basis for the non-invasive determination of the strength characteristics of the cerebral tissue aneurysms and determination of their status

A Trojan-horse peptide-carboxymethyl-cytidine antibiotic from Bacillus amyloliquefaciens

Microcin C and related antibiotics are Trojan-horse peptide-adenylates. The peptide part is responsible for facilitated transport inside the sensitive cell, where it gets processed to release a toxic warhead—a nonhydrolyzable aspartyl-adenylate, which inhibits aspartyl-tRNA synthetase. Adenylation of peptide precursors is carried out by MccB THIF-type NAD/FAD adenylyltransferases. Here, we describe a novel microcin C-like compound from Bacillus amyloliquefaciens. The B. amyloliquefaciens MccB demonstrates an unprecedented ability to attach a terminal cytidine monophosphate to cognate precursor peptide in cellular and cell free systems. The cytosine moiety undergoes an additional modification—carboxymethylation—that is carried out by the C-terminal domain of MccB and the MccS enzyme that produces carboxy-SAM, which serves as a donor of the carboxymethyl group. We show that microcin C-like compounds carrying terminal cytosines are biologically active and target aspartyl-tRNA synthetase, and that the carboxymethyl group prevents resistance that can occur due to modification of the warhead. The results expand the repertoire of known enzymatic modifications of peptides that can be used to obtain new biological activities while avoiding or limiting bacterial resistance.status: publishe