Oxidation Of Sulfur-containing Amino Acids Research Articles

Alkyl chain regulated cytomembrane and mitochondria targeting fluorescent probes for ratiometric sensing SO2 and its bio-application in mice model

Sulfur dioxide has been recognized as an important signaling molecule which can be generated enzymatically in mitochondria during oxidation of sulfur-containing amino acids. Meanwhile, cell membrane plays an important role in signal transduction. Therefore, the study of SO2 effects has proven to be challenging due to the lack of reliable tools for monitoring SO2 in different subcellular organelle. Herein, we designed two probes CPA-1 and CPA-2 based on coumarin derivatives. The cyano substituted double bond of the two probes were selected as the SO2 response site, and the length of the alkyl side chain of the pyridine salt was used to regulate the hydrophobicity to achieve the efficient targeting of the mitochondria and the cell membrane, respectively. Importantly, CPA-1 and CPA-2 shows ratiometric fluorescence response towards SO2 with highly sensitive and selective. Moreover, cell-imaging demonstrated that CPA-1 and CPA-2 possesses good mitochondria-targeted and cell membrane-targeted ability, respectively. Additionally, CPA-1 and CPA-2 were perfectly used for detection of SO2 in mice. We expect that the work will provide a useful tool and strategy for studying and visualizing signal molecules on the cell membrane in vivo.

Early Events of Photosensitized Oxidation of Sulfur-Containing Amino Acids Studied by Laser Flash Photolysis and Mass Spectrometry.

The mechanism of photooxidation of methionine (N-Ac-Met-NH-CH3, 1) and methyl-cysteine (N-Ac-MeCys-NH-CH3, 2) analogues by 3-carboxybenzophenone triplet (3CB*) in neutral aqueous solution was studied using techniques of nanosecond laser flash photolysis and steady-state photolysis. The short-lived transients derived from 3CB and sulfur-containing amino acids were identified, and their quantum yields and kinetics of formation and decay were determined. The stable photoproducts were analyzed using liquid chromatography coupled with high-resolution mass spectrometry. Substantial differences in the mechanisms were found for methionine and S-methyl-cysteine analogues for both primary and secondary photoreactions. A new secondary reaction channel (back hydrogen atom transfer from the ketyl radical to the carbon-centered α-thioalkyl radical yielding reactants in the ground states) was suggested. The detailed mechanisms of 3CB* sensitized photooxidation of 1 and 2 are proposed and discussed.

Селективное вольтамперометрическое определение серусодержащих аминокислот в лекарственных средствах и витаминных комплексах на электроде, модифицированном пленкой из гексахлороплатината рутения

Неорганическая пленка из гексахлороплатината рутения (III) (RuPtCl6), осажденная на поверхности электрода из стеклоуглерода, проявляет каталитическую активность при окислении серусодержащих аминокислот. Каталитическое электроокисление цистеина, цистина и метионина происходит при различных потенциалах. Выявлены оптимальные условия получения пленки RuPtCl6, при которых регистрируется наибольший каталитический эффект. Разработан способ селективного вольтамперометрического определения цистеина, метионина и цистина на электроде с пленкой RuPtCl6. Нижняя граница определяемых содержаний цистеина и цистина составляет 5 · 10–6 М, а метионина – 5 · 10–7 М. Предложенный способ вольтамперометрического определения использован для определения цистеина, цистина и метионина в лекарственных средствах и витаминных комплексах.

A dielectric barrier discharge terminally inactivates RNase A by oxidizing sulfur-containing amino acids and breaking structural disulfide bonds

RNases are among the most stable proteins in nature. They even refold spontaneously after heat inactivation, regaining full activity. Due to their stability and universal presence, they often pose a problem when experimenting with RNA. We investigated the capabilities of nonthermal atmospheric-pressure plasmas to inactivate RNase A and studied the inactivation mechanism on a molecular level.While prolonged heating above 90 °C is required for heat inactivating RNase A, direct plasma treatment with a dielectric barrier discharge (DBD) source caused permanent inactivation within minutes. Circular dichroism spectroscopy showed that DBD-treated RNase A unfolds rapidly. Raman spectroscopy indicated methionine modifications and formation of sulfonic acid. A mass spectrometry-based analysis of the protein modifications that occur during plasma treatment over time revealed that methionine sulfoxide formation coincides with protein inactivation. Chemical reduction of methionine sulfoxides partially restored RNase A activity confirming that sulfoxidation is causal and sufficient for RNase A inactivation. Continued plasma exposure led to over-oxidation of structural disulfide bonds. Using antibodies, disulfide bond over-oxidation was shown to be a general protein inactivation mechanism of the DBD. The antibody’s heavy and light chains linked by disulfide bonds dissociated after plasma exposure. Based on their ability to inactivate proteins by oxidation of sulfur-containing amino acids and over-oxidation of disulfide bonds, DBD devices present a viable option for inactivating undesired or hazardous proteins on heat or solvent-sensitive surfaces.

Redox Biology Poster Prize Winner P55 - Role of Oxidized Protein Repair in Human Skeletal Muscle

There is now increasing evidence that reactive oxygen species (ROS) are signalling molecules that regulate growth, differentiation, proliferation and apoptosis, at least in physiological concentration. However, when ROS levels overcome the capacity of cellular antioxidant systems, they damage cellular components such as nucleic acids, lipids and in particular proteins, inflicting alterations to cell structure and function. Oxidation of sulfur-containing aminoacids, like cysteine and methionine, within proteins, can be repaired by specific enzymatic systems. Indeed methionine sulfoxide is catalytically reduced back to methionine by the methionine sulfoxide reductase (Msr) system. We showed that this Msr system is involved in cellular protection against oxidative stress by preventing apoptosis and limiting irreversible protein oxidative damage. Furthermore, it has been demonstrated that the Msr system is no longer efficient during ageing and senescence as well as that Msr modulates longevity in animal models. In this work we analysed Msr expression in human skeletal muscle. We showed, using both primary and immortalized skeletal muscle cell lines, that the expression of the Msr system increases during differentiation, suggesting a role of this antioxidant system on myofibres formation. Since myoblasts senescence associated with less differentiation capacity and an abnormal production of ROS have been observed in some muscular dystrophies, we wondered about the state of the Msr system in late-onset myopathies, such as oculopharyngeal muscular dystrophy (OPMD). In OPMD we confirmed, at the transcription level, an increased expression of Msr with differentiation, both on immortalized and primary cell cultures, similarly to what we observed on control cell lines. Our aim is to characterize for the first time the role of a major oxidized protein repair system (Msr) in human skeletal muscle homeostasis and, in particular, in myotubes protection during oxidative stress conditions.

Effects of the Effluent of a Microscale Atmospheric Pressure Plasma-jet Operated with He/O2 Gas on Bovine Serum Albumin

Cold atmospheric pressure plasmas (CAPs) are being investigated for medical applications, and the first clinical studies are promising. However, interactions between plas- mas and biological samples are only partly understood on a molecular level. In this study, bo- vine serum albumin (BSA), a standard model for plasma-mediated etching of biological sam- ples, was used to investigate the effects of different components of an He/O 2 plasma effluent on proteins. The X-jet features an optional lateral helium flow that splits the plasma effluent into particles and (V)UV radiation. BSA samples were exposed separately to plasma-emitted particles, UV radiation, or the combination of both. Afterward, plasma-treated samples were investigated using SDS-PAGE and western blot analysis for amino acid strand breaks, but none were detected. Furthermore, treated samples were investigated by Raman spectroscopy to search for chemical modifications. We found that treatment with the X-jet has little effect on BSA. Minor changes in the Raman spectra suggest modifications of tyrosine residues and some degree of oxidation of sulfur-containing amino acids. Our findings suggest that for the X-jet effluent, etching is the main effect of plasma on BSA, making BSA a suitable model with which to study protein etching.

Virucidal mechanism of action of NVC-422, a novel antimicrobial drug for the treatment of adenoviral conjunctivitis

Human adenoviral conjunctivitis is a highly contagious eye infection affecting millions of people world-wide. If untreated, it can further develop into keratitis, corneal ulceration, scarring and possible blindness. Despite the significant patient morbidity and socio-economic costs, it is an unmet medical need with no FDA approved treatment. Here, we demonstrate the virucidal activity of NVC-422 (N,N-dichloro-2,2-dimethyltaurine) against adenovirus type 5 (Ad5) and investigated its mechanism of action of Ad5 inactivation. NVC-422 inhibits Ad5-induced loss of cell viability in vitro with 50% inhibitory concentration (IC50) ranging from 9 to 23μM. NVC-422 does not cause any cytotoxicity at concentrations as high as 250μM. Invitro, NVC-422 inactivates Ad5 but does not interfere with viral replication, indicating that NVC-422 acts on the extracellular adenovirus as a virucidal agent. NVC-422 inactivates Ad5 by oxidative inactivation of key viral proteins such as fiber and hexon as evidenced by SDS–PAGE, Western blotting and reversed-phase HPLC. These data, combined with measurements of the kinetics of the NVC-422 reactivity with selected amino acids, indicate that the changes in the viral proteins are caused by the selective oxidation of sulfur-containing amino acids. The conformational changes of the viral proteins result in the destruction of the viral morphology as shown by transmission electron microscopy. In summary, NVC-422 exhibits virucidal activity against Ad5 by the oxidative inactivation of key viral proteins, leading to the loss of viral integrity and infectivity.

Electrochemical oxidation of sulfur-containing amino acids on an electrode modified with multi-walled carbon nanotubes

MWNT-modified electrodes are introduced for the voltammetric determination of sulfur-containing amino acids. The morphology of the electrode surface has been characterized by atomic force microscopy. The MWNT layer consists of deeply intertwined vermicular structures with the average diameter of 25 nm. Cysteine, glutathione and methionine are oxidized on the electrode while only cysteine gives signals on the glassy carbon (GC) electrode. The application of such electrodes leads to a decreased overpotential and increase of oxidation currents for cysteine in comparison with a bare GC electrode. The schemes of oxidation are proposed. A decrease of the lower limit of determination and an enlargement of the analytical range for antioxidants were obtained. A simple, fast and accurate procedure for the voltammetric determination of methionine in pharmaceuticals has been developed and can be recommended for quality control purposes.

Methionine Sulfoxide Reduction and the Aging Process

Aging has been described for multicellular and asymmetrically dividing organisms, but the mechanisms are poorly understood. Oxidation of proteins is considered to be one of the major factors that leads to aging. Oxidative damage to proteins results in the oxidation of certain amino acid residues, among which oxidation of sulfur-containing amino acids, methionine and cysteine, is notable because of the susceptibility of these residues to damage, and occurrence of repair mechanisms. Methionine sulfoxide reductases, MsrA and MsrB, are thioredoxin-dependent oxidoreductases that reduce oxidized forms of methionine, methionine sulfoxides, in a stereospecific manner. These enzymes are present in all cell types and have shown to be regulating life spans in mammals, insects, and yeast. Here, their roles in modulating yeast life span are discussed.

First Steps in the Oxidation of Sulfur-Containing Amino Acids by Hypohalogenation: Very Fast Generation of Intermediate Sulfenyl Halides and Halosulfonium Cations

Sulfur-containing amino acids show an extraordinary binding towards HOCl/ClO −. During the process, the Cl is transferred from the O to the S of the amino acid. Met reacts with HOCl one order of magnitude faster than the non- S containing amino acids (k (Met+HOCl) =8.7·10 8 mol −1 dm 3 s −1). Instead, Cys reacts as its thiolate (RS −), two orders-of-magnitude faster (k ( RS −+ HOCl) =1.2·10 9 mol −1 dm 3 s −1). Cys reacts also with ClO − (k ( RS −+ ClO −) =1.9·10 5 mol −1 dm 3 s −1). Such processes take place much more readily than the corresponding N-halogenation of the non-sulfur containing amino acids. To our knowledge, these are the first kinetic measurements of the rate of formation of sulfenyl halides and halosulfonium cations in aqueous solution. Sulfenyl chlorides and chlorosulfonium ions derived from amino acids are elusive, and sulfide-type amino acids (Met) eventually yield sulfoxides (MetO), while thiol-type amino acids (Cys) lead to disulfides (Cys^Cys) and sulfonic acids (Cya). The fate of sulfur-containing amino acids upon oxidation with HOCl/ClO − seems to be related to their mutagen-inactivation ability.

Amperometric Detection of Biologically Significant Sulfur-Containing Compounds at Bi(V)-Doped PbO2Film Electrodes

Electrocatalytic oxidation of sulfur-containing amino acids and peptides at Bi(V)-doped β-PbO2 film electrodes (Bi-PbO2) was investigated in acidic media and tested for amperometric detection of representatives of these compounds following their separation as cationic species in a chromatographic column that exhibits both cation-exchange and reversed-phase properties. The Bi-PbO2 film electrodes permit detection with similar sensitivities for compounds that contain sulfhydryl, disulfide, thioether, and sulfoxide moieties. More specifically, detection limits at the pmol level were obtained for all analytes of interest. The Bi-PbO2 electrodes were characterized by cyclic voltammetry, scanning electron microscopy (SEM), and energy dispersive spectroscopy (EDS). For the acidic media used in this study, only the nonsulfur containing amino acids tryptophane and tyrosine produced an anodic response; however, this occurred only at concentrations >50 μM. The linear dynamic range of response for methionine is demonstrated to be a sensitive function of applied potential. This observation is concluded to be consistent with a response mechanism by which a prerequisite is the anodic discharge of H2O to produce adsorbed OH radicals that are the immediate source of O-atoms transferred to the oxidation products.

Excited Triplet State of N-(9-Methylpurin-6-yl)pyridinium Cation as an Efficient Photosensitizer in the Oxidation of Sulfur-Containing Amino Acids. Laser Flash and Steady-State Photolysis Studies [Erratum to document cited in CA122:81956

Excited Triplet State of N-(9-Methylpurin-6-yl)pyridinium Cation as an Efficient Photosensitizer in the Oxidation of Sulfur-Containing Amino Acids. Laser Flash and Steady-State Photolysis Studies [Erratum to document cited in CA122:81956

Excited Triplet State of N-(9-Methylpurin-6-yl)pyridinium Cation as an Efficient Photosensitizer in the Oxidation of Sulfur-Containing Amino Acids. Laser Flash and Steady-State Photolysis Studies

In this paper, we present and discuss the results of a detailed mechanistic study of the photosensitized oxidation of sulfur-containing amino acids by N-(9-methylpurin-6-yl)pyridinium cations in aqueous solution. The results provide data on the photoreactivity of the pyridinium salts (derived from the purine bases) in the presence of the electron donors such as sulfur-containing organic compounds and give new experimental evidence for the general mechanism of the sensitized photooxidation of thioethers proposed previously. 39 refs., 6 figs., 2 tabs.

The selective oxidation of sulfur-containing amino acids by diethyl azodicarboxylate.

The selective oxidation of sulfur-containing amino acids by diethyl azodicarboxylate.