The mechanism of action of nitro-heterocyclic antimicrobial drugs. Primary target of 1-methyl-2-nitro-5-vinylimidazole is DNA.

The antimicrobial drug 1-methyl-2-nitro-5-vinylimidazole (MEV) preferentially blocked DNA synthesis, was mutagenic and induced coliphage lambda in Escherichia coli. The antibacterial effects of MEV are the consequences of repairable damage to DNA, as shown by hypersensitivity of recA and uvr strains to MEV and related drugs, stimulation by MEV of DNA turnover which was dependent on the product of the uvrA gene, and the presence of cross-links in DNA from MEV-treated bacteria.     

The metabolism of aromatic acids by micro-organisms. Metabolic pathways in the fungi

1. The metabolic pathways of aromatic-ring fission were examined in a range of fungal genera that utilize several compounds related to lignin. 2. Most of the genera, after growth on p-hydroxybenzoate, protocatechuate or compounds that are degraded to the latter (e.g. caffeate, ferulate or vanillate), rapidly oxidized these compounds, but not catechol. 3. Such genera possessed a protocatechuate 3,4-oxygenase and accumulated beta-carboxymuconate as the product of protocatechuate oxidation. This enzyme had a high pH optimum in most organisms; the Rhodotorula enzyme was competitively inhibited by catechol. 4. beta-Carboxymuconate was converted by all competent fungi into beta-carboxymuconolactone, which was isolated and characterized. None of the fungi produced or utilized at significant rates the corresponding bacterial intermediate gamma-carboxymuconolactone. 5. The lactonizing enzymes of Rhodotorula and Neurospora crassa had a pH optimum near 5.5 and approximate molecular weights of 19000 and 190000 respectively. 6. The fungi did not degrade the isomeric (+)-muconolactone, gamma-carboxymethylenebutanolide or beta-oxoadipate enol lactone at significant rates, and thus differ radically from bacteria, where beta-oxoadipate enol lactone is the precursor of beta-oxoadipate in all strains examined. 7. The end product of beta-carboxymuconolactone metabolism by extracts was beta-oxoadipate. 8. Evidence for a coenzyme A derivative of beta-oxoadipate was found during further metabolism of this keto acid. 9. A few anomalous fungi, after growth on p-hydroxybenzoate, had no protocatechuate 3,4-oxygenase, but possessed all the enzymes of the catechol pathway. Catechol was detected in the growth medium in one instance. 10. A strain of Penicillium sp. formed pyruvate but no beta-oxoadipate from protocatechuate, suggesting the existence also of a ;meta' type of ring cleavage among fungi.     

Molecular mechanism of the antimicrobial action of pyocyanin

The mechanism by which pyocyanin inhibits bacterial growth was investigated. Several organisms possessing varying levels of superoxide dismutase were analyzed for their sensitivity to pyocyanin to test the possibility that reduced pyocyanin univalently reduces oxygen to superoxide, thus causing cell death. No correlation was found between the amount of superoxide dismutase possessed by an organism and resistance to pyocyanin. In addition, it was demonstrated that organisms growing anaerobically with nitrate as a terminal electron acceptor were as sensitive as, or more sensitive to the action of pyocyanin than organisms grown under aerobic conditions. We thus rule out the possibility that excess superoxide generation is the primary mechanism by which pyocyanin exerts its antibiotic effect. Oxygen electrode and radioisotope studies demonstrated that pyocyanin does inhibit bacterial respiration and active transport of solutes. Thus, it was concluded that the mechanism of action is the result of pyocyanin interacting with the cell membrane respiratory chain in such a way to render the cell unable to perform energy-requiring, membrane-bound metabolic processes such as active transport.     

On the mechanism of reductive activation in the mode of action of some anticancer drugs

Bioactivation of a number of DNA-specific antitumor drugs depends on oxidoreduction. Bleomycin, neocarzinostatin and anthracycline glycosides are the best known among such drugs in terms of reductive activation processes. Their reduction results in short-lived radical or electrophilic intermediates attacking DNA stereospecifically. The physico-chemical properties of these drugs and the nature of DNA damage are reviewed. Models for DNA-intercalation, electron-donor systems involved in drug metabolisation, and the role of oxygen in radical reactions, are discussed in the light of recent reports.     

香兰素抗菌机制研究进展

食品安全问题在中国范围内频发,严重威胁民众的健康和安全。为防止此类问题发生,对于由微生物污染食品所产生的危害,可通过添加抗菌剂有效降低安全隐患,随着食品安全法的不断完善,抗菌剂的添加量也愈加严格和规范。香兰素是一种具有抗菌性的传统食品添加剂,但是目前缺乏对其抗菌机制的全面理解,因此,限制了香兰素在抗菌特性方面的广泛应用。有鉴于此,现对香兰素的结构、功能与抗菌活性的相关性及其抗菌机制研究进展予以综述,并对具有抗菌特性的香兰素衍生物进行探讨。     

Elucidation of antimicrobial activity and mechanism of action by N-substituted carbazole derivatives

Compounds belonging to a carbazole series have been identified as potent fungal plasma membrane proton adenosine triphophatase (H⁺-ATPase) inhibitors with a broad spectrum of antifungal activity. The carbazole compounds inhibit the adenosine triphosphate (ATP) hydrolysis activity of the essential fungal H⁺-ATPase, thereby functionally inhibiting the extrusion of protons and extracellular acidification, processes that are responsible for maintaining high plasma membrane potential. The compound class binds to and inhibits the H⁺-ATPase within minutes, leading to fungal death after 1–3 h of compound exposure in vitro. The tested compounds are not selective for the fungal H⁺-ATPase, exhibiting an overlap of inhibitory activity with the mammalian protein family of P-type ATPases; the sarco(endo)plasmic reticulum calcium ATPase (Ca²⁺-ATPase) and the sodium potassium ATPase (Na⁺,K⁺-ATPase). The ion transport in the P-type ATPases is energized by the conversion of ATP to adenosine diphosphate (ADP) and phosphate and a general inhibitory mechanism mediated by the carbazole derivative could therefore be blocking of the active site. However, biochemical studies show that increased concentrations of ATP do not change the inhibitory activity of the carbazoles suggesting they act as allosteric inhibitors. Furthermore decreased levels of intracellular ATP would suggest that the compounds inhibit the H⁺-ATPase indirectly, but Candida albicans cells exposed to potent H⁺-ATPase-inhibitory carbazoles result in increased levels of intracellular ATP, indicating direct inhibition of H⁺-ATPase.     

Pyrrolomycins as antimicrobial agents. Microwave-assisted organic synthesis and insights into their antimicrobial mechanism of action

New compounds able to counteract staphylococcal biofilm formation are needed. In this study we investigate the mechanism of action of pyrrolomycins, whose potential as antimicrobial agents has been demonstrated. We performed a new efficient and easy method to use microwave organic synthesis suitable for obtaining pyrrolomycins in good yields and in suitable amount for their in vitro in-depth investigation. We evaluate the inhibitory activity towards Sortase A (SrtA), a transpeptidase responsible for covalent anchoring in Gram-positive peptidoglycan of many surface proteins involved in adhesion and in biofilm formation. All compounds show a good inhibitory activity toward SrtA, having IC₅₀ values ranging from 130 to 300?µM comparable to berberine hydrochloride. Of note compound 1d shows a good affinity in docking experiment to SrtA and exhibits the highest capability to interfere with biofilm formation of S. aureus showing an IC₅₀ of 3.4?nM. This compound is also effective in altering S. aureus murein hydrolase activity that is known to be responsible for degradation, turnover, and maturation of bacterial peptidoglycan and involved in the initial stages of S. aureus biofilm formation.     

The mechanism of action of dipeptidyl aminopeptidase. Inhibition by amino acid derivatives and amines; activation by aromatic compounds.

A variety of amino acid and peptide amides have been shown to be inhibitors of dipeptidyl aminopeptidase. Among these compounds derivatives of strongly hydrophobic amino acids are the strongest inhibitors (Phe-NH2, Ki = 1.0 +/- 0.2 mM), while amides of basic amino acids were somewhat less effective (Lys-NH2, Ki = 36 +/- 3 mM). Short chain amino acid amides are notably weaker inhibitors (Gly-NH2, Ki = 293 +/- 50 mM). The interaction of the side chains of compounds with the enzyme appears to be at a site other than that at which the side chain of the amino-penultimate residue of the substrate interacts since the specificity of binding is different. Primary amines have been shown to inhibit, e.g., butylamine, Ki = 340 +/- 40 mM, and aromatic compounds have been shown to stimulate activity toward Gly-Gly-NH2 and Gly-Gly-OEt (phenol, 35% stimulation of activity at a 1:1 molar ratio with the substrate). The data suggest that inhibition involves binding at the site occupied by the free alpha-amino group and the N-terminal amino acid.     

The mechanism of activation of rabbit plasminogen by urokinase.

The data presented in this paper show that when rabbit plasminogen is activated to plasmin by urokinase at least two peptide bonds are cleaved in the process. Urokinase first cleaves an internal peptide bond in plasminogen, leading to two-chain disulfide-linked plasmin molecule. The plasmin heavy chain of molecular weight 66,000 to 69,000 possesses an NH2-terminal amino acid sequence identical with the original plasminogen (molecular weight 88,000 to 92,000). The plasmin light chain of molecular weight 24,000 to 26,000 is known to be derived from the COOH-terminal portion of plasminogen. The plasmin generated during the activation of plasminogen is capable, by a feedback process, of cleaving a peptide of molecular weight 6,000 to 8,000 from the NH2 terminus of the heavy chain, producing a proteolytically modified heavy chain of molecular weight 58,000 to 62,000. Plasmin also can cleave this same peptide from the original plasminogen, yielding an altered plasminogen of molecular weight 82,000 to 86,000. This plasmin-altered plasminogen and the plasmin heavy chain derived from it by urokinase activation process NH2-terminal amino acid sequences which are identical with each other and with the plasminolytic product of the original plasmin heavy chain. These studies support a mechanism of activation of plasminogen by urokinase which involves loss of a peptide located on the NH2 terminus of plasminogen. However, these same results show that this NH2-terminal peptide need not be released from rabbit plasminogen prior to the cleavage of the internal peptide bond which leads to the two-chain plasmin molecule. Furthermore, these studies show that urokinase cannot remove this peptide from either the original rabbit plasminogen molecule or from the heavy chain of the initial plasmin formed.     

Structure and mechanism of action of the antimicrobial peptide piscidin

Piscidin, an antibacterial peptide isolated from the mast cells of striped bass, has potent antimicrobial activity against a broad spectrum of pathogens in vitro. We investigated the mechanism of action of this 22-residue cationic peptide by carrying out structural studies and electrophysiological experiments in lipid bilayers. Circular dichroism experiments showed that piscidin was unstructured in water but had a high alpha-helix content in dodecylphosphocholine (DPC) micelles. 1H NMR data in water and TFE confirmed these results and demonstrated that the segment of residues 8-17 adopted an alpha-helical structure in a micellar environment. This molecule has a marked amphipathic character, due to well-defined hydrophobic and hydrophilic sectors. This structure is similar to those determined for other cationic peptides involved in permeabilization of the bacterial membrane. Multichannel experiments with piscidin incorporated into azolectin planar bilayers gave reproducible I-V curves at various peptide concentrations and unambiguously showed that this peptide permeabilized the membrane. This pore forming activity was confirmed by single-channel experiments, with well-defined ion channels obtained at different voltages. The characteristics of the ion channels (voltage dependence, only one or two states of conductance) clearly suggest that piscidin is more likely to permeabilize the membrane by toroidal pore formation rather than via the "barrel-stave" mechanism.     

Lipid headgroup discrimination by antimicrobial peptide LL-37: insight into mechanism of action

Interaction of the human antimicrobial peptide LL-37 with lipid monolayers has been investigated by a range of complementary techniques including pressure-area isotherms, insertion assay, epifluorescence microscopy, and synchrotron x-ray scattering, to analyze its mechanism of action. Lipid monolayers were formed at the air-liquid interface to mimic the surface of the bacterial cell wall and the outer leaflet of erythrocyte cell membrane by using phosphatidylglycerol (DPPG), phosphatidylcholine (DPPC), and phosphatidylethanolamine (DPPE) lipids. LL-37 is found to readily insert into DPPG monolayers, disrupting their structure and thus indicating bactericidal action. In contrast, DPPC and DPPE monolayers remained virtually unaffected by LL-37, demonstrating its nonhemolytic activity and lipid discrimination. Specular x-ray reflectivity data yielded considerable differences in layer thickness and electron-density profile after addition of the peptide to DPPG monolayers, but little change was seen after peptide injection when probing monolayers composed of DPPC and DPPE. Grazing incidence x-ray diffraction demonstrated significant peptide insertion and lateral packing order disruption of the DPPG monolayer by LL-37 insertion. Epifluorescence microscopy data support these findings.