Biology subjects

Zamakhaeva, S.

Publications and source records attributed to Zamakhaeva, S..

5 recordsLinked to original sources

bioRxiv2026

Enterococcus faecalis autolysin, EpaU, binds the enterococcal polysaccharide antigen via its teichoic acid-like repeats and modulates c-di-AMP signaling

The cell wall of the Gram-positive bacterium Enterococcus faecalis is decorated with the enterococcal polysaccharide antigen (EPA), consisting of a core rhamnan backbone linked covalently with a strain-variable teichoic acid-like (TA) polymer. Current models propose that the TA decoration is a repeating polymer composed of two alternating subunits, designated TAI and TAII, which are attached to the rhamnan core via a mild-acid labile phosphodiester bond from the initiating TAI subunit. In this study, we characterize the EpaU autolysin encoded within the EPA biosynthetic gene cluster. We demonstrate that the cell wall-binding domain of EpaU associates with the intact TA domains of EPA synthesized with the aid of the glycosyltransferases EpaR and EpaX. We further show that EpaU is a potent autolysin that binds generally over the E. faecalis cell surface, suggesting that it functions as a remodeling peptidoglycan hydrolase. The absence of EpaU leads to increased ampicillin resistance and elevated intracellular levels of the second messenger c-di-AMP. These data suggest that E. faecalis possesses a mechanism that senses the integrity of the peptidoglycan meshwork and employs c-di-AMP to regulate cell turgor, potentially altering the antibiotic resistance. ImportanceEnterococcus faecalis is an important opportunistic pathogen that can cause severe nosocomial infections. Knowledge of how bacteria remodel the cell wall is key to understanding many important cellular processes, such as antibiotic resistance, cell division, biofilm formation, and stress resistance. In this study, we shed new light on the structural details of the main cell wall polysaccharide, Enterococcal Polysaccharide Antigen (EPA), and its interaction with EpaU, an autolysin that cleaves peptidoglycan during cell wall remodeling. We also report a link between EpaU and the regulation of turgor pressure via cyclic dinucleotide signaling. This work contributes to a more complete picture of E. faecalis cell wall and may provide insight into the development of antimicrobial agents based on autolysins.

microbiology↗

bioRxiv2024

Structure and mechanism of biosynthesis of Streptococcus mutans cell wall polysaccharide

Streptococcus mutans, the causative agent of human dental caries, expresses a cell wall attached Serotype c-specific Carbohydrate (SCC) that is critical for cell viability. SCC consists of a repeating [->]3)-Rha(1[->]2)-Rha(1[->] polyrhamnose backbone, with glucose (Glc) side-chains and glycerol phosphate (GroP) decorations. This study reveals that SCC has one major and two minor Glc modifications. The major Glc modification, -Glc, attached to position 2 of 3-rhamnose, is installed by SccN and SccM glycosyltransferases and is the site of the GroP addition. The minor Glc modifications are {beta}-Glc linked to position 4 of 3-rhamnose installed by SccP and SccQ glycosyltransferases, and -Glc attached to position 4 of 2-rhamnose installed by SccN working in tandem with an unknown enzyme. Both the major and the minor {beta}-Glc modifications control bacterial morphology, but only the GroP and major Glc modifications are critical for biofilm formation.

microbiology↗

bioRxiv2024

Disorder regulates homeostasis of extracytoplasmic proteins in streptococci

Proteins harboring intrinsically disordered regions (IDRs) lacking stable secondary or tertiary structures are abundant across the three domains of life. These regions have not been systematically studied in prokaryotes. Our genome-wide analysis identifies extracytoplasmic serine/threonine-rich IDRs in several biologically important membrane-associated proteins in streptococci. We demonstrate that these IDRs are glycosylated with glucose by glycosyltransferases GtrB and PgtC2 in Streptococcus pyogenes and Streptococcus pneumoniae, and with N-acetylgalactosamine by a Pgf-dependent mechanism in Streptococcus mutans. The absence of glycosylation leads to a defect in biofilm formation under ethanol-stressed conditions in S. mutans. We link this phenotype to the C-terminal IDR of the post-translocation chaperone PrsA. Our data reveal that O-linked glycosylation protects the IDR-containing proteins from proteolytic degradation and is critical for the biological function of PrsA in biofilm formation.

microbiology↗

bioRxiv2021

PplD is a de-N-acetylase of the cell wall linkage unit of streptococcal rhamnopolysaccharides

The cell wall of the human bacterial pathogen Group A Streptococcus (GAS) consists of peptidoglycan decorated with the Lancefield group A carbohydrate (GAC). GAC is a promising target for the development of GAS vaccines. In this study, employing chemical, compositional, and NMR methods, we show that GAC is attached to peptidoglycan via glucosamine 1-phosphate. This structural feature makes the GAC-peptidoglycan linkage highly sensitive to cleavage by nitrous acid and resistant to mild acid conditions. Using this characteristic of the GAS cell wall, we identify PplD as a protein required for deacetylation of linkage N-acetylglucosamine (GlcNAc). X-ray structural analysis indicates that PplD performs catalysis via a modified acid/base mechanism. Genetic surveys in silico together with functional analysis indicate that PplD homologs deacetylate the polysaccharide linkage in many streptococcal species. We further demonstrate that introduction of positive charges to the cell wall by GlcNAc deacetylation protects GAS against host cationic antimicrobial proteins.

microbiology↗

bioRxiv2020

Modification of streptococcal cell wall polysaccharide spatially controls cell division

Bacterial cell division is driven by a tubulin homolog FtsZ, which assembles into the Z-ring structure leading to the recruitment of the cell division machinery. In ovoid-shaped Gram-positive bacteria, such as streptococci, MapZ guides Z-ring positioning at cell equators through an, as yet, unknown mechanism. The cell wall of the important dental pathogen Streptococcus mutans is composed of peptidoglycan decorated with Serotype c Carbohydrates (SCCs). Here, we show that an immature form of SCC, lacking the recently identified glycerol phosphate (GroP) modification, coordinates Z-ring positioning. Pulldown assays using S. mutans cell wall combined with binding affinity analysis identified the major cell separation autolysin, AtlA, as an SCC binding protein. Importantly, AtlA binding to mature SCC is attenuated due to GroP modification. Using fluorescently-labeled AtlA, we mapped SCC distribution on the streptococcal surface to reveal that GroP-deficient immature SCCs are exclusively present at the cell poles and equators. Moreover, the equatorial GroP-deficient SCCs co-localize with MapZ throughout the S. mutans cell cycle. Consequently, in GroP-deficient mutant bacteria, proper AtlA localization is abrogated resulting in dysregulated cellular autolysis. In addition, these mutants display morphological abnormalities associated with MapZ mislocalization leading to Z-ring misplacement. Altogether, our data support a model in which GroP-deficient immature SCCs spatially coordinate the localization of AtlA and MapZ. This mechanism ensures cell separation by AtlA at poles and Z-ring alignment with the cell equator.Competing Interest StatementThe authors have declared no competing interest.View Full Text

microbiology↗