sep413

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Structural basis for selective inhibition of antibacterial target MraY, a membrane-bound enzyme involved in peptidoglycan synthesis.

Drug Discov Today. 2018 May 17;:

Authors: Hering J, Dunevall E, Ek M, Brändén G

Abstract
The rapid growth of antibiotic-resistant bacterial infections is of major concern for human health. Therefore, it is of great importance to characterize novel targets for the development of antibacterial drugs. One promising protein target is MraY (UDP-N-acetylmuramyl-pentapeptide: undecaprenyl phosphate N-acetylmuramyl-pentapeptide-1-phosphate transferase or MurNAc-1-P-transferase), which is essential for bacterial cell wall synthesis. Here, we summarize recent breakthroughs in structural studies of bacterial MraYs and the closely related human GPT (UDP-N-acetylglucosamine: dolichyl phosphate N-acetylglucosamine-1-phosphate transferase or GlcNAc-1-P-transferase). We present a detailed comparison of interaction modes with the natural product inhibitors tunicamycin and muraymycin D2. Finally, we speculate on possible routes to design an antibacterial agent in the form of a potent and selective inhibitor against MraY.

PMID: 29778697 [PubMed - as supplied by publisher]

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Whole genome sequencing data of 1110 Mycobacterium tuberculosis isolates identifies insertions and deletions associated with drug resistance.

BMC Genomics. 2018 May 16;19(1):365

Authors: Zeng X, Kwok JS, Yang KY, Leung KS, Shi M, Yang Z, Yam WC, Tsui SK

Abstract
BACKGROUND: Drug resistance in Mycobacterium tuberculosis (MTB) is one of the major challenges in tuberculosis (TB) treatment. However, known mutations cannot explain all of the cases of resistance and little research has focused on the relationship between insertions / deletions (indels) and drug resistance.
RESULTS: Here, we retrieved whole genome sequencing data of 743 drug-resistant MTB strains and 367 pan-susceptible strains from TB patients from the public domain to identify novel genomic markers of drug resistance. A total of 20 region markers containing genes and intergenic regions (IGRs) with significant statistical correlation with antibiotic resistance were revealed, four of which have been previously reported to be associated with drug resistance. In addition, 83 point markers containing frameshift (FS) mutations and IGR indels were also identified independently based on differences in their incidence rates between drug-sensitive and -resistant strains. Among the 83 point markers, eight indels were detected in known drug-associated genes or IGRs. Furthermore, the overlap between 20 region markers and 83 point markers further indicated their associations with drug resistance. The markers identified were involved in essential bacterial metabolic functions, including cell wall and transmembrane transporter functions. A strong correlation between FS mutations and mutations in DNA repair genes including I21V in alkA, R48G in mutT4 and P2R in nth was also found.
CONCLUSIONS: This study identified a set of novel genetic markers with FS mutations and IGR indels associated with MTB drug resistance, which greatly broadens the pool of mutations related to MTB drug resistance. This insight may be important in identifying novel mechanisms of drug resistance in MTB.

PMID: 29769016 [PubMed - in process]

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Multiple Click-Selective tRNA Synthetases Expand Mammalian Cell-Specific Proteomics.

J Am Chem Soc. 2018 06 13;140(23):7046-7051

Authors: Yang AC, du Bois H, Olsson N, Gate D, Lehallier B, Berdnik D, Brewer KD, Bertozzi CR, Elias JE, Wyss-Coray T

Abstract
Bioorthogonal tools enable cell-type-specific proteomics, a prerequisite to understanding biological processes in multicellular organisms. Here we report two engineered aminoacyl-tRNA synthetases for mammalian bioorthogonal labeling: a tyrosyl ( ScTyrY43G) and a phenylalanyl ( MmPheT413G) tRNA synthetase that incorporate azide-bearing noncanonical amino acids specifically into the nascent proteomes of host cells. Azide-labeled proteins are chemoselectively tagged via azide-alkyne cycloadditions with fluorophores for imaging or affinity resins for mass spectrometric characterization. Both mutant synthetases label human, hamster, and mouse cell line proteins and selectively activate their azido-bearing amino acids over 10-fold above the canonical. ScTyrY43G and MmPheT413G label overlapping but distinct proteomes in human cell lines, with broader proteome coverage upon their coexpression. In mice, ScTyrY43G and MmPheT413G label the melanoma tumor proteome and plasma secretome. This work furnishes new tools for mammalian residue-specific bioorthogonal chemistry, and enables more robust and comprehensive cell-type-specific proteomics in live mammals.

PMID: 29775058 [PubMed - indexed for MEDLINE]

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Computation-Guided Rational Design of a Peptide Motif That Reacts with Cyanobenzothiazoles via Internal Cysteine-Lysine Relay.

J Org Chem. 2018 07 20;83(14):7467-7479

Authors: Keyser SGL, Utz A, Bertozzi CR

Abstract
Site-selective protein modification based on covalent reactions of peptide tags and small molecules is a key capability for basic research as well as for the development of new therapeutic bioconjugates. Here, we describe the computation-guided rational design of a cysteine- and lysine-containing 11-residue peptide sequence that reacts with 2-cyanobenzothiazole (CBT) derivatives. Our data show that the cysteine residue reversibly reacts with the nitrile group on the CBT moiety to form an intermediate thioimidate, which undergoes irreversible SN transfer to the lysine residue, yielding an amidine-linked product. The concepts outlined herein lay a foundation for future development of peptide tags in the context of site-selective modification of lysine residues within engineered microenvironments.

PMID: 29771122 [PubMed - indexed for MEDLINE]

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Structure and Function of the Transmembrane Domain of NsaS, an Antibiotic Sensing Histidine Kinase in Staphylococcus aureus.

J Am Chem Soc. 2018 06 20;140(24):7471-7485

Authors: Bhate MP, Lemmin T, Kuenze G, Mensa B, Ganguly S, Peters JM, Schmidt N, Pelton JG, Gross CA, Meiler J, DeGrado WF

Abstract
NsaS is one of four intramembrane histidine kinases (HKs) in Staphylococcus aureus that mediate the pathogen's response to membrane active antimicrobials and human innate immunity. We describe the first integrative structural study of NsaS using a combination of solution state NMR spectroscopy, chemical-cross-linking, molecular modeling and dynamics. Three key structural features emerge: First, NsaS has a short N-terminal amphiphilic helix that anchors its transmembrane (TM) bundle into the inner leaflet of the membrane such that it might sense neighboring proteins or membrane deformations. Second, the transmembrane domain of NsaS is a 4-helix bundle with significant dynamics and structural deformations at the membrane interface. Third, the intracellular linker connecting the TM domain to the cytoplasmic catalytic domains of NsaS is a marginally stable helical dimer, with one state likely to be a coiled-coil. Data from chemical shifts, heteronuclear NOE, H/D exchange measurements and molecular modeling suggest that this linker might adopt different conformations during antibiotic induced signaling.

PMID: 29771498 [PubMed - indexed for MEDLINE]

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The cell wall hydrolase Pmp23 is important for assembly and stability of the division ring in Streptococcus pneumoniae.

Sci Rep. 2018 05 15;8(1):7591

Authors: Jacq M, Arthaud C, Manuse S, Mercy C, Bellard L, Peters K, Gallet B, Galindo J, Doan T, Vollmer W, Brun YV, VanNieuwenhze MS, Di Guilmi AM, Vernet T, Grangeasse C, Morlot C

Abstract
Bacterial division is intimately linked to synthesis and remodeling of the peptidoglycan, a cage-like polymer that surrounds the bacterial cell, providing shape and mechanical resistance. The bacterial division machinery, which is scaffolded by the cytoskeleton protein FtsZ, includes proteins with enzymatic, structural or regulatory functions. These proteins establish a complex network of transient functional and/or physical interactions which preserve cell shape and cell integrity. Cell wall hydrolases required for peptidoglycan remodeling are major contributors to this mechanism. Consistent with this, their deletion or depletion often results in morphological and/or division defects. However, the exact function of most of them remains elusive. In this work, we show that the putative lysozyme activity of the cell wall hydrolase Pmp23 is important for proper morphology and cell division in the opportunistic human pathogen Streptococcus pneumoniae. Our data indicate that active Pmp23 is required for proper localization of the Z-ring and the FtsZ-positioning protein MapZ. In addition, Pmp23 localizes to the division site and interacts directly with the essential peptidoglycan synthase PBP2x. Altogether, our data reveal a new regulatory function for peptidoglycan hydrolases.

PMID: 29765094 [PubMed - indexed for MEDLINE]

Researchers find a link between molecules released by commensal bacteria and the degree of inflammation in a mouse model of MS.

An inhibitor of starch-digesting enzymes stunts the growth of certain gut bacteria

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Cross-specificity of protective human antibodies against Klebsiella pneumoniae LPS O-antigen.

Nat Immunol. 2018 May 14;:

Authors: Rollenske T, Szijarto V, Lukasiewicz J, Guachalla LM, Stojkovic K, Hartl K, Stulik L, Kocher S, Lasitschka F, Al-Saeedi M, Schröder-Braunstein J, von Frankenberg M, Gaebelein G, Hoffmann P, Klein S, Heeg K, Nagy E, Nagy G, Wardemann H

Abstract
Humoral immune responses to microbial polysaccharide surface antigens can prevent bacterial infection but are typically strain specific and fail to mediate broad protection against different serotypes. Here we describe a panel of affinity-matured monoclonal human antibodies from peripheral blood immunoglobulin M-positive (IgM+) and IgA+ memory B cells and clonally related intestinal plasmablasts, directed against the lipopolysaccharide (LPS) O-antigen of Klebsiella pneumoniae, an opportunistic pathogen and major cause of antibiotic-resistant nosocomial infections. The antibodies showed distinct patterns of in vivo cross-specificity and protection against different clinically relevant K. pneumoniae serotypes. However, cross-specificity was not limited to K. pneumoniae, as K. pneumoniae-specific antibodies recognized diverse intestinal microbes and neutralized not only K. pneumoniae LPS but also non-K. pneumoniae LPS. Our data suggest that the recognition of minimal glycan epitopes abundantly expressed on microbial surfaces might serve as an efficient humoral immunological mechanism to control invading pathogens and the large diversity of the human microbiota with a limited set of cross-specific antibodies.

PMID: 29760533 [PubMed - as supplied by publisher]

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New strategies for targeting and treatment of multi-drug resistant Staphylococcus aureus.

Drug Resist Updat. 2017 Mar;31:1-14

Authors: Assis LM, Nedeljković M, Dessen A

Abstract
Staphylococcus aureus is a major cause of bacterial infection in humans, and has been notoriously able to acquire resistance to a variety of antibiotics. An example is methicillin-resistant S. aureus (MRSA), which despite having been initially associated with clinical settings, now is one of the key causative agents of community-acquired infections. Antibiotic resistance in S. aureus involves mechanisms ranging from drug efflux to increased expression or mutation of target proteins, and this has required innovative approaches to develop novel treatment methodologies. This review provides an overview of the major mechanisms of antibiotic resistance developed by S. aureus, and describes the emerging alternatives being sought to circumvent infection and proliferation, including new generations of classic antibiotics, synergistic approaches, antibodies, and targeting of virulence factors.

PMID: 28867240 [PubMed - indexed for MEDLINE]

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Spatial control of cell envelope biosynthesis in mycobacteria.

Pathog Dis. 2018 Jun 01;76(4):

Authors: Puffal J, García-Heredia A, Rahlwes KC, Siegrist MS, Morita YS

Abstract
The mycobacterial cell envelope is a complex multilayered structure that provides the strength to the rod-shaped cell and creates the permeability barrier against antibiotics and host immune attack. In this review, we will discuss the spatial coordination of cell envelope biosynthesis and how plasma membrane compartmentalization plays a role in this process. The spatial organization of cell envelope biosynthetic enzymes as well as other membrane-associated proteins is crucial for cellular processes such as polar growth and midcell septum formation. We will highlight metabolic enzymes involved in the localized biosynthesis of envelope components such as peptidoglycan, arabinogalactan and outer/inner membrane lipids. The known and potential roles of cytoskeletal and coiled coil proteins in driving subcellular protein localization will also be summarized. Finally, we provide a comprehensive overview of known lateral heterogeneities in mycobacterial plasma membrane, with a particular focus on the intracellular membrane domain, recently revealed by biochemical fractionation and fluorescence microscopy. We consider how this dynamic and multifunctional membrane microdomain contributes to the subcellular localization of membrane proteins and spatially restricted cell envelope biosynthesis in mycobacteria.

PMID: 29762679 [PubMed - in process]

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Site-specific incorporation of quadricyclane into a protein and photocleavage of the quadricyclane ligation adduct.

Bioorg Med Chem. 2018 10 15;26(19):5280-5290

Authors: Tomlin FM, Gordon CG, Han Y, Wu TS, Sletten EM, Bertozzi CR

Abstract
The quadricyclane (QC) ligation is a bioorthogonal reaction between a quadricyclane moiety and a nickel bis(dithiolene) derivative. Here we show that a QC amino acid can be incorporated into a protein site-specifically using the pyrrolysine-based genetic code expansion platform, and subsequently used for ligation chemistry. Additionally, we exploited the photolability of the QC ligation product to render the adduct cleavable with a handheld UV lamp. We further developed a protein purification method that involves QC ligation of biotin to a protein of interest, capture on streptavidin resin, and finally release using only UV light. The QC ligation thus brings novel chemical manipulations to the realm of bioorthogonal chemistry.

PMID: 29754834 [PubMed - indexed for MEDLINE]

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Glyco-seek: Ultrasensitive Detection of Protein-Specific Glycosylation by Proximity Ligation Polymerase Chain Reaction.

J Am Chem Soc. 2016 08 31;138(34):10722-5

Authors: Robinson PV, Tsai CT, de Groot AE, McKechnie JL, Bertozzi CR

Abstract
We report a non-destructive biochemical technique, termed "Glyco-seek", for analysis of O-GlcNAcylated proteins. Glyco-seek combines chemoenzymatic labeling, proximity ligation, and quantitative polymerase chain reaction to detect O-GlcNAcylated proteins with ultrahigh sensitivity. Our glycan-specific assay can be paired with traditional proximity ligation assays to simultaneously determine the change in total protein levels. We show that Glyco-seek detects attomoles of glycoproteins of interest from cell lysates, with sensitivity several orders of magnitude higher than that of current techniques. We used the method to directly assay the O-GlcNAcylation status of a low-abundance transcription factor from cell lysates without need for isolation or enrichment.

PMID: 27454154 [PubMed - indexed for MEDLINE]

Publication date: August 2018
Source:Biochimica et Biophysica Acta (BBA) - Biomembranes, Volume 1860, Issue 8
Author(s): Qiu Ying Lau, Jianguo Li, Marc-Antoine Sani, Sheetal Sinha, Yan Li, Fui Mee Ng, CongBao Kang, Surajit Bhattacharjya, Frances Separovic, Chandra Verma, Cheng San Brian Chia
Linear antimicrobial peptides, with their rapid bactericidal mode of action, are well-suited for development as topical antibacterial drugs. We recently designed a synthetic linear 4-residue peptide, BRBR-NH2, with potent bactericidal activity against Staphylococcus aureus (MIC 6.25 μM), the main causative pathogen of human skin infections with an unknown mechanism of action. Herein, we describe a series of experiments conducted to gain further insights into its mechanism of action involving electron microscopy, artificial membrane dye leakage, solution- and solid-state NMR spectroscopy followed by molecular dynamics simulations. Experimental results point towards a SMART (Soft Membranes Adapt and Respond, also Transiently) mechanism of action, suggesting that the peptide can be developed as a topical antibacterial agent for treating drug-resistant Staphylococcus aureus infections.

Graphical abstract

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Novobiocin Enhances Polymyxin Activity by Stimulating Lipopolysaccharide Transport.

J Am Chem Soc. 2018 06 06;140(22):6749-6753

Authors: Mandler MD, Baidin V, Lee J, Pahil KS, Owens TW, Kahne D

Abstract
Gram-negative bacteria are challenging to kill with antibiotics due to their impenetrable outer membrane containing lipopolysaccharide (LPS). The polymyxins, including colistin, are the drugs of last resort for treating Gram-negative infections. These drugs bind LPS and disrupt the outer membrane; however, their toxicity limits their usefulness. Polymyxin has been shown to synergize with many antibiotics including novobiocin, which inhibits DNA gyrase, by facilitating transport of these antibiotics across the outer membrane. Recently, we have shown that novobiocin not only inhibits DNA gyrase but also binds and stimulates LptB, the ATPase that powers LPS transport. Here, we report the synthesis of novobiocin derivatives that separate these two activities. One analog retains LptB-stimulatory activity but is unable to inhibit DNA gyrase. This analog, which is not toxic on its own, nevertheless enhances the lethality of polymyxin by binding LptB and stimulating LPS transport. Therefore, LPS transport agonism contributes substantially to novobiocin-polymyxin synergy. We also report other novobiocin analogs that inhibit DNA gyrase better than or equal to novobiocin, but bind better to LptB and therefore have even greater LptB stimulatory activity. These compounds are more potent than novobiocin when used in combination with polymyxin. Novobiocin analogs optimized for both gyrase inhibition and LPS transport agonism may allow the use of lower doses of polymyxin, increasing its efficacy and safety.

PMID: 29746111 [PubMed - indexed for MEDLINE]

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Diversity of Growth Patterns Probed in Live Cyanobacterial Cells Using a Fluorescent Analog of a Peptidoglycan Precursor.

Front Microbiol. 2018;9:791

Authors: Zhang JY, Lin GM, Xing WY, Zhang CC

Abstract
Cyanobacteria were the first oxygenic photosynthetic organisms during evolution and were ancestors of plastids. Cyanobacterial cells exhibit an extraordinary diversity in their size and shape, and bacterial cell morphology largely depends on the synthesis and the dynamics of the peptidoglycan (PG) layer. Here, we used a fluorescence analog of the PG synthesis precursor D-Ala, 7-Hydroxycoumarin-amino-D-alanine (HADA), to probe the PG synthesis pattern in live cells of cyanobacteria with different morphology. They displayed diverse synthesis patterns, with some strains showing an intensive HADA incorporation at the septal region, whereas others gave an HADA signal distributed around the cells. Growth zones covering several cells at the tips of the filament were present in some filamentous strains such as in Arthrospira. In Anabaena PCC 7120, which is capable of differentiating heterocysts for N2 fixation, PG synthesis followed the cell division cycle. In addition, an HADA incorporation was strongly activated from 12 to 15 h following the initiation of heterocyst development, indicating a thickening of the PG layer in heterocysts. The PG synthesis pattern is diverse in cyanobacteria and responds to developmental regulation. The use of fluorescent analogs may serve as a useful tool for understanding the mechanisms of cell growth and morphogenesis operating in these organisms.

PMID: 29740419 [PubMed]

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A novel whole-cell biosensor of Pseudomonas aeruginosa to monitor the expression of quorum sensing genes.

Appl Microbiol Biotechnol. 2018 Jul;102(14):6023-6038

Authors: Zhang C, Parrello D, Brown PJB, Wall JD, Hu Z

Abstract
A novel whole-cell biosensor was developed to noninvasively and simultaneously monitor the in situ genetic activities of the four quorum sensing (QS) networks in Pseudomonas aeruginosa PAO1, including the las, rhl, pqs, and iqs systems. P. aeruginosa PAO1 is a model bacterium for studies of biofilm and pathogenesis while both processes are closely controlled by the QS systems. This biosensor worked well by selectively monitoring the expression of one representative gene from each network. In the biosensor, the promoter regions of lasI, rhlI, pqsA, and ambB (QS genes) controlled the fluorescent reporter genes of Turbo YFP, mTag BFP2, mNEON Green, and E2-Orange, respectively. The biosensor was successful in monitoring the impact of an important environmental factor, salt stress, on the genetic regulation of QS networks. High salt concentrations (≥ 20 g·L-1) significantly downregulated rhlI, pqsA, and ambB after the biosensor was incubated for 17 h to 18 h at 37 °C, resulting in slow bacterial growth.

PMID: 29730766 [PubMed - indexed for MEDLINE]

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Activity-Related Conformational Changes in d,d-Carboxypeptidases Revealed by In Vivo Periplasmic Förster Resonance Energy Transfer Assay in Escherichia coli.

MBio. 2017 Sep 12;8(5):

Authors: Meiresonne NY, van der Ploeg R, Hink MA, den Blaauwen T

Abstract
One of the mechanisms of β-lactam antibiotic resistance requires the activity of d,d-carboxypeptidases (d,d-CPases) involved in peptidoglycan (PG) synthesis, making them putative targets for new antibiotic development. The activity of PG-synthesizing enzymes is often correlated with their association with other proteins. The PG layer is maintained in the periplasm between the two membranes of the Gram-negative cell envelope. Because no methods existed to detect in vivo interactions in this compartment, we have developed and validated a Förster resonance energy transfer assay. Using the fluorescent-protein donor-acceptor pair mNeonGreen-mCherry, periplasmic protein interactions were detected in fixed and in living bacteria, in single samples or in plate reader 96-well format. We show that the d,d-CPases PBP5, PBP6a, and PBP6b of Escherichia coli change dimer conformation between resting and active states. Complementation studies and changes in localization suggest that these d,d-CPases are not redundant but that their balanced activity is required for robust PG synthesis.IMPORTANCE The periplasmic space between the outer and the inner membrane of Gram-negative bacteria contains many essential regulatory, transport, and cell wall-synthesizing and -hydrolyzing proteins. To date, no assay is available to determine protein interactions in this compartment. We have developed a periplasmic protein interaction assay for living and fixed bacteria in single samples or 96-well-plate format. Using this assay, we were able to demonstrate conformation changes related to the activity of proteins that could not have been detected by any other living-cell method available. The assay uniquely expands our toolbox for antibiotic screening and mode-of-action studies.

PMID: 28900026 [PubMed - indexed for MEDLINE]

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Two DD-carboxypeptidases from Mycobacterium smegmatis affect cell surface properties through regulation of peptidoglycan cross-linking and glycopeptidolipids.

J Bacteriol. 2018 May 07;:

Authors: Pandey SD, Pal S, Kumar N G, Bansal A, Mallick S, Ghosh AS

Abstract
During the peptidoglycan (PG) maturation of mycobacteria, the glycan strands are interlinked by both 3-3 (between two meso-DAP) and 4-3 cross-links (between D-ala and meso-DAP), though there is a predominance (60-80%) of 3-3 cross-links. The DD-CPases act on pentapeptides to generate tetrapeptides that are used by LD-transpeptidases as substrates to form 3-3 cross-links. Therefore, DD-CPases play a crucial role in mycobacterial PG cross-link formation. However, the physiology of DD-CPases in mycobacteria is relatively unexplored. Here, we deleted two DD-CPase genes, msmeg_2433, and msmeg_2432, both individually and in combination, from Mycobacterium smegmatis mc2155. Though the single DD-CPase deletions had no significant impact on the mycobacterial physiology, many interesting functional alterations were observed in the double deletion mutant, viz., a predominance in PG cross-link formation was shifted from 3-3 cross-links to 4-3, cell surface glycopeptidolipid (GPL) expression was reduced and susceptibility towards β-lactams and anti-tubercular agents was enhanced. Moreover, the existence of the double mutant within murine macrophages was better as compared to the parent. Interestingly, the complementation with any one of the DD-CPase genes could restore the wild-type phenotype. In a nutshell, we infer that the altered ratio of 4-3: 3-3 PG cross-links might have influenced the expression of surface GPLs, colony morphology, biofilm formation,, drug susceptibility and subsistence of the cells within macrophages.Importance The glycan strands in mycobacterial peptidoglycan (PG) are interlinked by both 3-3 and 4-3 cross-links. The DD-CPases generate tetrapeptides by acting on the pentapeptides, and LD-transpeptidases use tetrapeptides as substrates to form 3-3 cross-links. Here, we showed that simultaneous deletions of two DD-CPases alter the nature of PG cross-linking from 3-3 cross-links to 4-3 cross-links. The deletions subsequently decrease the expression of Glycopeptidolipids (a significant surface lipid present in many non-tuberculous mycobacteria including Mycobacterium smegmatis) and affect other physiological parameters like cell morphology, growth rate, biofilm formation, antibiotic susceptibility and existence within murine macrophages. Thus, unraveling the physiology of DD-CPases might help us design anti-mycobacterial therapeutics in future.

PMID: 29735762 [PubMed - as supplied by publisher]

New drugs are needed to treat gram-negative bacterial infections. These bacteria are protected by an outer membrane which prevents many antibiotics from reaching their cellular targets. The outer leaflet of the outer membrane contains LPS, which is responsible for creating this permeability barrier. Interfering with LPS biogenesis affects bacterial viability....

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Hidden Mode of Action of Glycopeptide Antibiotics: Inhibition of Wall Teichoic Acid Biosynthesis.

J Phys Chem B. 2017 04 27;121(16):3925-3932

Authors: Singh M, Chang J, Coffman L, Kim SJ

Abstract
Glycopeptide antibiotics inhibit the peptidoglycan biosynthesis in Gram-positive bacteria by targeting lipid II. This prevents the recycling of bactoprenol phosphate, the lipid transporter that is shared by peptidoglycan and wall teichoic acid biosyntheses. In this study, we investigate the effects of glycopeptide antibiotics on peptidoglycan and wall teichoic acid biosynthesis. The incorporation of d-[1-13C]alanine, d-[15N]alanine, and l-[1-13C]lysine into peptidoglycan and wall teichoic acid in intact whole cells of Staphylococcus aureus was measured using 13C{15N} and 15N{13C} rotational-echo double resonance NMR. S. aureus treated with oritavancin and vancomycin at subminimal inhibitory concentrations exhibit a large reduction in d-Ala incorporation into wall teichoic acid, but without changes to the peptidoglycan cross-links or the stem-links. Thus, sequestration of bactoprenol phosphate by glycopeptide antibiotics resulted in inhibition of d-Ala incorporation into the wall teichoic acid prior to the inhibition of peptidoglycan biosynthesis. Our finding shows that S. aureus responds to glycopeptide-induced cell wall stress by routing all available d-Ala to the peptidoglycan biosynthesis, at the cost of reducing the wall teichoic acid biosynthesis.

PMID: 28368603 [PubMed - indexed for MEDLINE]