Rachita Dash

J Antimicrob Chemother. 2022 Nov 28;77(12):3256-3264. doi: 10.1093/jac/dkac309.

ABSTRACT

BACKGROUND: Infections caused by bacterial biofilms are very difficult to treat. The use of currently approved antibiotics even at high dosages often fails, making the treatment of these infections very challenging. Novel antimicrobial agents that use distinct mechanisms of action are urgently needed.

OBJECTIVES: To explore the use of [G1K,K8R]cGm, a designed cyclic analogue of the antimicrobial peptide gomesin, as an alternative approach to treat biofilm infections.

METHODS: We studied the activity of [G1K,K8R]cGm against biofilms of Staphylococcus aureus, a pathogen associated with several biofilm-related infections. A combination of atomic force and real-time confocal laser scanning microscopies was used to study the mechanism of action of the peptide.

RESULTS: The peptide demonstrated potent activity against 24 h-preformed biofilms through a concentration-dependent ability to kill biofilm-embedded cells. Mechanistic studies showed that [G1K,K8R]cGm causes morphological changes on bacterial cells and permeabilizes their membranes across the biofilm with a half-time of 65 min. We also tested an analogue of [G1K,K8R]cGm without disulphide bonds, and a linear unfolded analogue, and found both to be inactive.

CONCLUSIONS: The results suggest that the 3D structure of [G1K,K8R]cGm and its stabilization by disulphide bonds are essential for its antibacterial and antibiofilm activities. Moreover, our findings support the potential application of this stable cyclic antimicrobial peptide to fight bacterial biofilms.

PMID:36171717 | PMC:PMC9704431 | DOI:10.1093/jac/dkac309

ACS Infect Dis. 2022 Oct 17. doi: 10.1021/acsinfecdis.2c00393. Online ahead of print.

ABSTRACT

Due to a steady increase in microbial resistance, there is a need to increase the effectiveness of antibiotic performance by involving additional mechanisms of their penetration or retention for their better action. Cephalosporins are a successful group of antibiotics to combat pathogenic microorganisms, including drug-resistant strains. In this study, we investigated the effect of newly synthesized cephalosporin derivatives with cyclic disulfide modifications against several Gram-positive and Gram-negative strains as well as against biofilm formation. The incorporation of asparagusic acid was found to be effective in improving the activity of the drug against Gram-negative strains compared to the all carbon-control compounds. Furthermore, we could demonstrate the successful reduction of biofilm formation for Staphylococcus aureus and Pseudomonas aeruginosa at similar concentrations as obtained against planktonic cells. We propose that the incorporation of cyclic disulfides is one additional strategy to improve antibiotic activity and to combat bacterial infections.

PMID:36251034 | DOI:10.1021/acsinfecdis.2c00393

ACS Infect Dis. 2022 Oct 18. doi: 10.1021/acsinfecdis.2c00406. Online ahead of print.

ABSTRACT

Antibiotics have been widely used in the medical field as a treatment for infectious diseases, but they are not effective against all Gram-negative bacteria because of their low permeability to the outer membrane. One of the strategies to improve the antibacterial activity of antibiotics is the coadministration of antibiotics and membrane-perturbing antimicrobial peptides for their synergistic effects. However, because of their different pharmacokinetics, their coadministration may not exert expected effects in the clinical stage. Here, we designed various antimicrobial peptide-antibiotic conjugates as a novel approach to improve the antimicrobial activity of antibiotics. Ampicillin was chosen as a model antibiotic with poor outer membrane permeability, and the effects of the chemistry and position of conjugation and the choice of antimicrobial peptides were examined. One of the ampicillin conjugates exhibited significantly improved antimicrobial activity against ampicillin-resistant Gram-negative bacteria without exerting cytotoxicity against human cultured cells, demonstrating that our novel approach is an effective strategy to improve the antimicrobial activity of antibiotics with low outer membrane permeability.

PMID:36255133 | DOI:10.1021/acsinfecdis.2c00406

Cell. 2022 Sep 20:S0092-8674(22)01191-6. doi: 10.1016/j.cell.2022.09.019. Online ahead of print.

ABSTRACT

The G protein-coupled receptor cascade leading to production of the second messenger cAMP is replete with pharmacologically targetable proteins, with the exception of the Gα subunit, Gαs. GTPases remain largely undruggable given the difficulty of displacing high-affinity guanine nucleotides and the lack of other drug binding sites. We explored a chemical library of 10¹² cyclic peptides to expand the chemical search for inhibitors of this enzyme class. We identified two macrocyclic peptides, GN13 and GD20, that antagonize the active and inactive states of Gαs, respectively. Both macrocyclic peptides fine-tune Gαs activity with high nucleotide-binding-state selectivity and G protein class-specificity. Co-crystal structures reveal that GN13 and GD20 distinguish the conformational differences within the switch II/α3 pocket. Cell-permeable analogs of GN13 and GD20 modulate Gαs/Gβγ signaling in cells through binding to crystallographically defined pockets. The discovery of cyclic peptide inhibitors targeting Gαs provides a path for further development of state-dependent GTPase inhibitors.

PMID:36170854 | DOI:10.1016/j.cell.2022.09.019

Adv Exp Med Biol. 2022;1386:147-184. doi: 10.1007/978-3-031-08491-1_6.

ABSTRACT

Bacteria sense their environment via the cell envelope, which in Gram-negative bacteria comprises the outer membrane, the periplasmic space, and the inner membrane. Pseudomonas aeruginosa is an opportunistic pathogen which is exposed to different cell wall stresses imposed by exposure to antibiotics, osmotic pressure, and long-time colonization of host tissues such as the lung in cystic fibrosis patients. In response to these stresses, P. aeruginosa is able to respond by establishing a cell envelope stress response involving different regulatory pathways including the extra-cytoplasmic sigma factors AlgU, SigX, and SbrI and other two-component sensor/response regulators and effectors. This chapter aims to review the different factors leading to the activation of the cell envelope stress response in P. aeruginosa and the genetic determinants involved in this response, which is crucial for the survival of the bacterium upon exposure to different stressful conditions.

PMID:36258072 | DOI:10.1007/978-3-031-08491-1_6

Chemistry. 2022 Oct 18. doi: 10.1002/chem.202202991. Online ahead of print.

ABSTRACT

Soluble fragments of peptidoglycan called muropeptides are released from the cell wall of bacteria as part of their metabolism or as a result of biological stresses. These compounds trigger immune responses in mammals and plants. In bacteria, they play a major role in the induction of antibiotic resistance. The development of efficient methods to produce muropeptides is therefore desirable both to address their mechanism of action and to design new antibacterial and immunostimulant agents. Here, we engineered the peptidoglycan recycling pathway of Escherichia coli to produce N-acetyl-β-D-glucosaminyl-(1→4)-1,6-anhydro-N-acetyl-β-D-muramic acid (GlcNAc-anhMurNAc), a common precursor of Gram-negative and Gram-positive muropeptides. Inactivation of the hexosaminidase nagZ gene allowed the efficient production of this key disaccharide, providing access to Gram-positive muropeptides through subsequent chemical peptide conjugation. E. coli strains deficient in both NagZ hexosaminidase and amidase activities further enabled the in vivo production of Gram-negative muropeptides containing meso-diaminopimelic acid, a rarely available amino acid.

PMID:36256497 | DOI:10.1002/chem.202202991

ACS Infect Dis. 2022 Aug 12;8(8):1627-1636. doi: 10.1021/acsinfecdis.2c00229. Epub 2022 Aug 2.

ABSTRACT

The rise of antibiotic-resistant Mycobacterium tuberculosis and non-tuberculous mycobacterial infections has placed ever-increasing importance on discovering new antibiotics to treat these diseases. Recently, a new penem, T405, was discovered to have strong antimicrobial activity against M. tuberculosis and Mycobacteroides abscessus. Here, a penem library of C2 side-chain variants was synthesized, and their antimicrobial activities were evaluated against M. tuberculosis H₃₇Rv and M. abscessus ATCC 19977. Several new penems with antimicrobial activity stronger than the standard-of-care carbapenem antibiotics were identified with some candidates improving on the activity of the lead compound, T405. Moreover, many candidates showed little or no increase in the minimum inhibitory concentration in the presence of serum compared to the highly protein-bound T405. The penems with the strongest activity identified in this study were then biochemically characterized by reaction with the representative l,d-transpeptidase Ldt_Mt2 and the representative penicillin-binding protein d,d-carboxypeptidase DacB2.

PMID:35916356 | PMC:PMC10029149 | DOI:10.1021/acsinfecdis.2c00229

Cancers (Basel). 2022 Oct 9;14(19):4945. doi: 10.3390/cancers14194945.

ABSTRACT

Owing to its unique mechanism of abundant pathogen-associated molecular patterns in antitumor immune responses, bacteria-based cancer immunotherapy has recently attracted wide attention. Compared to traditional cancer treatments such as surgery, chemotherapy, radiotherapy, and phototherapy, bacteria-based cancer immunotherapy exhibits the versatile capabilities for suppressing cancer thanks to its preferentially accumulating and proliferating within tumors. In particular, bacteria have demonstrated their anticancer effect through the toxins, and other active components from the cell membrane, cell wall, and dormant spores. More importantly, the design of engineering bacteria with detoxification and specificity is essential for the efficacy of bacteria-based cancer therapeutics. Meanwhile, bacteria can deliver the cytokines, antibody, and other anticancer theranostic nanoparticles to tumor microenvironments by regulating the expression of the bacterial genes or chemical and physical loading. In this review, we illustrate that naïve bacteria and their components can serve as robust theranostic agents for cancer eradication. In addition, we summarize the recent advances in efficient antitumor treatments by genetically engineering bacteria and bacteria-based nanoparticles. Further, possible future perspectives in bacteria-based cancer immunotherapy are also inspected.

PMID:36230868 | PMC:PMC9563255 | DOI:10.3390/cancers14194945

Molecules. 2022 Sep 29;27(19):6440. doi: 10.3390/molecules27196440.

ABSTRACT

The development of new techniques to rapidly and accurately detect bacteria has drawn continuous attention due to the potential threats posed by bacteria to human health and safety. Recently, a novel strategy based on fluorescent probes has drawn considerable interest for the detection of bacteria due to its high selectivity, fast response, and simple operation. In this review, we summarize the recent progress on fluorescent probes for the specific recognition and discrimination of Gram-negative and Gram-positive bacteria. In particular, we outline current design strategies, such as targeting of the differences in surface components, cell wall components, endogenous enzymes, surface charge, and hydrophobicity of various kinds of bacteria to develop various fluorescent sensors (organic small-molecule fluorescent probes, nanoprobes, and metal ion probes). We also emphasize the application of organic molecules in probe recognition elements. We hope that this review can stimulate this research area in bacterial detection and imaging in the future.

PMID:36234978 | PMC:PMC9572786 | DOI:10.3390/molecules27196440

Biochem Biophys Rep. 2022 Oct 8;32:101367. doi: 10.1016/j.bbrep.2022.101367. eCollection 2022 Dec.

ABSTRACT

The blood-brain barrier (BBB) is a major hurdle in drug discovery for central nervous system (CNS) disorders. Particularly, mid-size molecules and macromolecules (e.g., peptides and antibodies) that modulate intractable drug targets such as protein-protein interaction are prevented from entering the CNS via BBB. The receptor-mediated transcytosis (RMT) pathway has been examined to deliver these molecules to CNS. Among the receptors, low-density lipoprotein receptor-related protein 1 (LRP1) has been emerged as one of the promising receptors for RMT. Although several LRP1-binding peptides have been reported, no drugs are available on the market based on the combination of reported LRP1-binding peptides and therapeutic molecules. One reason may be stability in vivo and BBB-permeability of the peptides. The present study aims to identify a novel LRP1-binding peptide for RMT, where we successfully generated a 15-mer cyclic peptide named KS-487. It explicitly bound to Cluster 4 domain of LRP1 with the binding EC₅₀ value of 10.5 nM and was relatively stable in mouse plasma within 24 h. Moreover, its high BBB permeability was demonstrated using in vitro rat and monkey BBB models. By 24 h incubation, 13% and 17% of the added amount of KS-487 (10 μM) penetrated rat BBB and monkey BBB, respectively. KS-487 would be a potential candidate for the LRP1-mediated transcytosis-based drug delivery to CNS, as these values were significantly higher than those of the known LRP1-binding peptides-Angiopep-2 and L57.

PMID:36237444 | PMC:PMC9552116 | DOI:10.1016/j.bbrep.2022.101367

Gut Microbes. 2022 Jan-Dec;14(1):2125737. doi: 10.1080/19490976.2022.2125737.

ABSTRACT

Transmission of bacterial endospores between the environment and people and the following germination in vivo play critical roles in both the deadly infections of some bacterial pathogens and the stabilization of the commensal microbiotas in humans. Our knowledge about the germination process of different bacteria in the mammalian gut, however, is still very limited due to the lack of suitable tools to visually monitor this process. We proposed a two-step labeling strategy that can image and quantify the endospores' germination in the recipient's intestines. Endospores collected from donor's gut microbiota were first labeled with fluorescein isothiocyanate and transplanted to mice via gavage. The recipient mice were then administered with Cyanine5-tagged D-amino acid to label all the viable bacteria, including the germinated endospores, in their intestines in situ. The germinated donor endospores could be distinguished by presenting two types of fluorescent signals simultaneously. The integrative use of cell-sorting, 16S rDNA sequencing, and fluorescence in situ hybridization (FISH) staining of the two-colored bacteria unveiled the taxonomic information of the donor endospores that germinated in the recipient's gut. Using this strategy, we investigated effects of different germinants and pre-treatment interventions on their germination, and found that germination of different commensal bacterial genera was distinctly affected by various types of germinants. This two-color labeling strategy shows its potential as a versatile tool for visually monitoring endospore germination in the hosts and screening for new interventions to improve endospore-based therapeutics.

PMID:36175402 | PMC:PMC9543051 | DOI:10.1080/19490976.2022.2125737

Life (Basel). 2022 Aug 28;12(9):1333. doi: 10.3390/life12091333.

ABSTRACT

Tuberculosis (TB) remains one of the deadliest infectious diseases in the world. Although several established antitubercular drugs have been found, various factors obstruct efforts to combat this disease due to the existence of drug-resistance (DR) TB strains, the need for lengthy treatment, and the occurrence of side effects from drug-drug interactions. Rifampicin (RIF) is the first line of antitubercular drugs and targets RNA polymerase (RNAP) of Mycobacterium tuberculosis (MTB). Here, RIF blocks the synthesis of long RNA during transcription initiation. The efficacy of RIF is low in DR-TB strains, and the use of RIF leads to various side effects. In this study, novel cyclic peptides were computationally designed as inhibitors of MTB transcription initiation. The designed cyclic peptides were subjected to a virtual screening to generate compounds that can bind to the RIF binding site in MTB RNAP subunit β (RpoB) for obtaining a new potential TB drug with a safe clinical profile. The molecular simulations showed that the cyclic peptides were capable of binding with RpoB mutants, suggesting that they can be possibility utilized for treating DR-TB. Structural modifications were carried out by acetylation and amidation of the N- and C-terminus, respectively, to improve their plasma stability and bioavailability. The modified linear and cyclic peptides were successfully synthesized with a solid-phase peptide synthesis method using Fmoc chemistry, and they were characterized by analytical HPLC, LC-ESI-MS⁺, and 1H NMR.

PMID:36143370 | PMC:PMC9506182 | DOI:10.3390/life12091333

Pharmaceutics. 2022 Sep 15;14(9):1956. doi: 10.3390/pharmaceutics14091956.

ABSTRACT

Acyldepsipeptides (ADEPs) are a new class of emerging antimicrobial peptides (AMPs), which are currently explored for treatment of pathogenic infections, including tuberculosis (TB). These cyclic hydrophobic peptides have a unique bacterial target to the conventional anti-TB drugs, and present a therapeutic window to overcome Mycobacterium Tuberculosis (M. tb) drug resistance. ADEPs exerts their antibacterial activity on M. tb strains through activation of the protein homeostatic regulatory protease, the caseinolytic protease (ClpP1P2). ClpP1P2 is normally regulated and activated by the ClpP-ATPases to degrade misfolded and toxic peptides and/or short proteins. ADEPs bind and dysregulate all the homeostatic capabilities of ClpP1P2 while inducing non-selective proteolysis. The uncontrolled proteolysis leads to M. tb cell death within the host. ADEPs analogues that have been tested possess cytotoxicity and poor pharmacokinetic and pharmacodynamic properties. However, these can be improved by drug design techniques. Moreover, the use of nanomaterial in conjunction with ADEPs would yield effective synergistic effect. This new mode of action has potential to combat and eradicate the extensive multi-drug resistance (MDR) problem that is currently faced by the public health pertaining bacterial infections, especially TB.

PMID:36145704 | PMC:PMC9502522 | DOI:10.3390/pharmaceutics14091956

World J Gastroenterol. 2022 Aug 21;28(31):4471-4474. doi: 10.3748/wjg.v28.i31.4471.

ABSTRACT

Irritable bowel syndrome (IBS) is an important health care concern. Alterations in the microbiota of the gut-brain axis may be linked to the pathophysiology of IBS. Some dietary intake could contribute to produce various metabolites including D-amino acids by the fermentation by the gut microbiota. D-amino acids are the enantiomeric counterparts of L-amino acids, in general, which could play key roles in cellular physiological processes against various oxidative stresses. Therefore, the presence of D-amino acids has been shown to be linked to the protection of several organs in the body. In particular, the gut microbiota could play significant roles in the stability of emotion via the action of D-amino acids. Here, we would like to shed light on the roles of D-amino acids, which could be used for the treatment of IBS.

PMID:36159020 | PMC:PMC9453761 | DOI:10.3748/wjg.v28.i31.4471

Microbiol Spectr. 2022 Sep 19:e0299022. doi: 10.1128/spectrum.02990-22. Online ahead of print.

ABSTRACT

Bacterial efflux pumps in the resistance-nodulation-cell division (RND) family of Gram-negative bacteria contribute significantly to the development of antimicrobial resistance by many pathogens. In this study, we selected the MtrD transporter protein of Neisseria gonorrhoeae as it is the sole RND pump possessed by this strictly human pathogen and can export multiple antimicrobials, including antibiotics, bile salts, detergents, dyes, and antimicrobial peptides. Using knowledge from our previously published structures of MtrD in the presence or absence of bound antibiotics as a model and the known ability of MtrCDE to export cationic antimicrobial peptides, we hypothesized that cationic peptides could be accommodated within MtrD binding sites. Furthermore, we thought that MtrD-bound peptides lacking antibacterial action could sensitize bacteria to an antibiotic normally exported by the MtrCDE efflux pump or other similar RND-type pumps possessed by different Gram-negative bacteria. We now report the identification of a novel nonantimicrobial cyclic cationic antimicrobial peptide, which we termed CASP (cationic antibiotic-sensitizing peptide). By single-particle cryo-electron microscopy, we found that CASP binds within the periplasmic cleft region of MtrD using overlapping and distinct amino acid contact sites that interact with another cyclic peptide (colistin) or a linear human cationic antimicrobial peptide derived from human LL-37. While CASP could not sensitize Neisseria gonorrhoeae to an antibiotic (novobiocin) that is a substrate for RND pumps, it could do so against multiple Gram-negative, rod-shaped bacteria. We propose that CASP (or future derivatives) could serve as an adjuvant for the antibiotic treatment of certain Gram-negative infections previously thwarted by RND transporters. IMPORTANCE RND efflux pumps can export numerous antimicrobials that enter Gram-negative bacteria, and their action can reduce the efficacy of antibiotics and provide decreased susceptibility to various host antimicrobials. Here, we identified a cationic antibiotic-sensitizing peptide (CASP) that binds within the periplasmic cleft of an RND transporter protein (MtrD) produced by Neisseria gonorrhoeae. Surprisingly, CASP was able to render rod-shaped Gram-negative bacteria, but not gonococci, susceptible to an antibiotic that is a substrate for the gonococcal MtrCDE efflux pump. CASP (or its future derivatives) could be used as an adjuvant to treat infections for which RND efflux contributes to multidrug resistance.

PMID:36121287 | DOI:10.1128/spectrum.02990-22