The reaction of alkylating agents with bacteriophage R17. Biological effects of phosphotriester formation.
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ABSTRACT:
SUBMITTER: Shooter KV
PROVIDER: S-EPMC1166119 | biostudies-other | 1974 Feb
REPOSITORIES: biostudies-other
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Project description:1. The inactivation of an RNA-containing bacteriophage after reaction with four methylating agents was studied. Measurements of the extent of methylation of the RNA and of the nature and amounts of the various reaction products were made. In experiments with dimethyl sulphate and methyl methanesulphonate inactivation can be quantitatively accounted for by methylation at two of the positions involved in hydrogen bonding: N-1 of adenine and N-3 of cytosine. In experiments with N-methyl-N-nitrosourea and N-methyl-N'-nitro-N-nitrosoguanidine methylation at N-1 of adenine and N-3 of cytosine accounts for only about one-half of the observed inactivation. Scission of the RNA chain during reaction accounts for a further 20% of the inactivation. To account for the remainder it seems necessary to postulate that formation of O(6)-methylguanine constitutes a lethal lesion. 2. Breaks in the RNA chain formed on reaction with the nitroso derivatives presumably result from methylation of the phosphate diester group followed by hydrolysis of the unstable triester thus formed.
| S-EPMC1166118 | biostudies-other
Project description:Recent advances in therapies have yielded notable success in terms of improved survival in several cancers. However, such treatments have failed to improve outcome in patients with gliomas for whom surgery followed by radiation therapy and chemotherapy with alkylating agents remain the standard of care. Genetic and epigenetic studies have helped identify several alterations specific to gliomas. Attempts to target these altered pathways have been unsuccessful due to various factors, including tumor heterogeneity, adaptive resistance of tumor cells, and limitations of access across the blood-brain barrier. Novel therapies that circumvent such limitations have been the focus of intense study and include approaches such as immunotherapy, targeting of signaling hubs and metabolic pathways, and use of biologic agents. Immunotherapeutic approaches including tumor-targeted vaccines, immune checkpoint blockade, antibody-drug conjugates, and chimeric antigen receptor-expressing cell therapies are in various stages of clinical trials. Similarly, identification of key metabolic pathways or converging hubs of signaling pathways that are tumor specific have yielded novel targets for therapy of gliomas. In addition, the failure of conventional therapies against gliomas has led to a growing interest among patients in the use of alternative therapies, which in turn has necessitated developing evidence-based approaches to the application of such therapies in clinical studies. The development of these novel approaches bears potential for providing breakthroughs in treatment of more meaningful and improved outcomes for patients with gliomas.
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Project description:Animal models for retinal degeneration are essential for elucidating its pathogenesis and developing new therapeutic strategies in humans. N-methyl-N-nitrosourea (MNU) has been extensively used to construct a photoreceptor-specific degeneration model, which has served to unveil the molecular process of photoreceptor degeneration as well as the mechanisms regulating the protective responses of remaining cells. Methyl methanesulphonate (MMS), also known to cause photoreceptor degeneration, is considered a good alternative to MNU due to its higher usability; however, detailed pathophysiological processes after MMS treatment remain uncharacterized. Here, we analyzed the time course of photoreceptor degeneration, Müller glial proliferation, and expression of secretory factors after MNU and MMS treatments in rats. While the timing of rod degeneration was similar between the treatments, we unexpectedly found that cones survived slightly longer after MMS treatment. Müller glia reentered the cell cycle at a similar timing after the two treatments; however, the G1/S transition occurred earlier after MMS treatment. Moreover, growth factors such as FGF2 and LIF were more highly upregulated in the MMS model. These data suggest that comparative analyses of the two injury models may be beneficial for understanding the complex regulatory mechanisms underlying the proliferative response of Müller glia.
| S-EPMC10762013 | biostudies-literature
Project description:Bacteria of the genus Staphylococcus are significant challenge for medicine, as many species are resistant to multiple antibiotics and some are even to all of the antibiotics we use. One of the approaches to developing new therapeutics to treat staphylococcal infections is the use of bacteriophages specific to these bacteria or the lytic enzymes of such bacteriophages, which are capable of hydrolyzing the cell walls of these bacteria. In this study, a new bacteriophage vB_SepP_134 (St 134) specific to Staphylococcus epidermidis was described. This podophage, with a genome of 18,275 bp, belongs to the Andhravirus genus. St 134 was able to infect various strains of 12 of the 21 tested coagulase-negative Staphylococcus species and one clinical strain from the Staphylococcus aureus complex. The genes encoding endolysin (LysSte134_1) and tail tip lysin (LysSte134_2) were identified in the St 134 genome. Both enzymes were cloned and produced in Escherichia coli cells. The endolysin LysSte134_1 demonstrated catalytic activity against peptidoglycans isolated from S. aureus, S. epidermidis, Staphylococcus haemolyticus, and Staphylococcus warneri. LysSte134_1 was active against S. aureus and S. epidermidis planktonic cells and destroyed the biofilms formed by clinical strains of S. aureus and S. epidermidis.
| S-EPMC10975630 | biostudies-literature
Project description:BackgroundThe appearance of plaques on a bacterial lawn is one of the enduring imageries in modern day biology. The seeming simplicity of a plaque has invited many hypotheses and models in trying to describe and explain the details of its formation. However, until now, there has been no systematic experimental exploration on how different bacteriophage (phage) traits may influence the formation of a plaque. In this study, we constructed a series of isogenic ? phages that differ in their adsorption rate, lysis timing, or morphology so that we can determine the effects if these changes on three plaque properties: size, progeny productivity, and phage concentration within plaques.ResultsWe found that the adsorption rate has a diminishing, but negative impact on all three plaque measurements. Interestingly, there exists a concave relationship between the lysis time and plaque size, resulting in an apparent optimal lysis time that maximizes the plaque size. Although suggestive in appearance, we did not detect a significant effect of lysis time on plaque productivity. Nonetheless, the combined effects of plaque size and productivity resulted in an apparent convex relationship between the lysis time and phage concentration within plaques. Lastly, we found that virion morphology also affected plaque size. We compared our results to the available models on plaque size and productivity. For the models in their current forms, a few of them can capture the qualitative aspects of our results, but not consistently in both plaque properties.ConclusionsBy using a collection of isogenic phage strains, we were able to investigate the effects of individual phage traits on plaque size, plaque productivity, and average phage concentration in a plaque while holding all other traits constant. The controlled nature of our study allowed us to test several model predictions on plaque size and plaque productivity. It seems that a more realistic theoretical approach to plaque formation is needed in order to capture the complex interaction between phage and its bacterium host in a spatially restricted environment.
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Project description: Not available
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Project description:Biological molecular assemblies are excellent models for the development of nanoengineered systems with desirable biomedical properties. Here we report an approach for fabrication of spontaneous, biologically active molecular networks consisting of bacteriophage (phage) directly assembled with gold (Au) nanoparticles (termed Au-phage). We show that when the phage are engineered so that each phage particle displays a peptide, such networks preserve the cell surface receptor binding and internalization attributes of the displayed peptide. The spontaneous organization of these targeted networks can be manipulated further by incorporation of imidazole (Au-phage-imid), which induces changes in fractal structure and near-infrared optical properties. The networks can be used as labels for enhanced fluorescence and dark-field microscopy, surface-enhanced Raman scattering detection, and near-infrared photon-to-heat conversion. Together, the physical and biological features within these targeted networks offer convenient multifunctional integration within a single entity with potential for nanotechnology-based biomedical applications.
| S-EPMC1346765 | biostudies-literature
Project description:Redox processes and radical intermediates are found in many biochemical processes, including deoxyribonucleotide synthesis and oxidative DNA damage. One of the core principles underlying DNA biosynthesis is the radical-mediated elimination of H2O to deoxygenate ribonucleotides, an example of 'spin-centre shift', during which an alcohol C-O bond is cleaved, resulting in a carbon-centred radical intermediate. Although spin-centre shift is a well-understood biochemical process, it is underused by the synthetic organic chemistry community. We wondered whether it would be possible to take advantage of this naturally occurring process to accomplish mild, non-traditional alkylation reactions using alcohols as radical precursors. Because conventional radical-based alkylation methods require the use of stoichiometric oxidants, increased temperatures or peroxides, a mild protocol using simple and abundant alkylating agents would have considerable use in the synthesis of diversely functionalized pharmacophores. Here we describe the development of a dual catalytic alkylation of heteroarenes, using alcohols as mild alkylating reagents. This method represents the first, to our knowledge, broadly applicable use of unactivated alcohols as latent alkylating reagents, achieved via the successful merger of photoredox and hydrogen atom transfer catalysis. The value of this multi-catalytic protocol has been demonstrated through the late-stage functionalization of the medicinal agents, fasudil and milrinone.
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Project description:Derivatives of methyl 3-(1-methyl-5-(1-methyl-5-(propylcarbamoyl)-1H-pyrrol-3-ylcarbamoyl)-1H-pyrrol-3-ylamino)-3-oxopropane-1-sulfonate (1), a peptide-based DNA minor groove binding methylating agent, were synthesized and characterized. In all cases, the N-terminus was appended with an O-methyl sulfonate ester, while the C-terminus group was varied with nonpolar and polar side chains. In addition, the number of pyrrole rings was varied from 2 (dipeptide) to 3 (tripeptide). The ability of the different analogues to efficiently generate N3-methyladenine was demonstrated as was their selectivity for minor groove (N3-methyladenine) versus major groove (N7-methylguanine) methylation. Induced circular dichroism studies were used to measure the DNA equilibrium binding properties of the stable sulfone analogues; the tripeptide binds with affinity that is >10-fold higher than that of the dipeptide. The toxicities of the compounds were evaluated in alkA/tag glycosylase mutant E. coli and in human WT glioma cells and in cells overexpressing and under-expressing N-methylpurine-DNA glycosylase, which excises N3-methyladenine from DNA. The results show that equilibrium binding correlates with the levels of N3-methyladenine produced and cellular toxicity. The toxicity of 1 was inversely related to the expression of MPG in both the bacterial and mammalian cell lines. The enhanced toxicity parallels the reduced activation of PARP and the diminished rate of formation of aldehyde reactive sites observed in the MPG knockdown cells. It is proposed that unrepaired N3-methyladenine is toxic due to its ability to directly block DNA polymerization.
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Project description:DNA polymerase iota (Pol?) belongs to the Y-family of DNA polymerases that are involved in DNA damage tolerance through their role in translesion DNA synthesis. Like all other Y-family polymerases, Pol? interacts with proliferating cell nuclear antigen (PCNA), Rev1, ubiquitin and ubiquitinated-PCNA and is also ubiquitinated itself. Here, we report that Pol? also interacts with the p300 acetyltransferase and is acetylated. The primary acetylation site is K550, located in the Rev1-interacting region. However, K550 amino acid substitutions have no effect on Pol?'s ability to interact with Rev1. Interestingly, we find that acetylation of Pol? significantly and specifically increases in response to SN2 alkylating agents and to a lower extent to SN1 alkylating and oxidative agents. As we have not observed acetylation of Pol?'s closest paralogue, DNA polymerase eta (Pol?), with which Pol? shares many functional similarities, we believe that this modification might exclusively regulate yet to be determined, and separate function(s) of Pol?.
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