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1.
Biomed Khim ; 66(1): 18-29, 2020 Jan.
Artigo em Russo | MEDLINE | ID: mdl-32116223

RESUMO

Mass spectrometric proteomic analysis at the sample preparation stage involves the artificial reduction of disulfide bonds in proteins formed between cysteine residues. Such bonds, when preserved in their native state, complicate subsequent enzymatic hydrolysis and interpretation of the research results. To prevent the re-formation of the disulfide bonds, cysteine residues are protected by special groups, most often by alkylation. In this review, we consider the methods used to modify cysteine residues during sample preparation, as well as possible artifacts of this stage. Particularly, adverse reactions of the alkylating agents with other amino acid residues are described. The most common alkylating compound used to protect cysteine residues in mass spectrometric proteomic analysis is iodoacetamide. However, an analysis of the literature in this area indicates that this reagent causes more adverse reactions than other agents used, such as chloroacetamide and acrylamide. The latter can be recommended for wider use. In the review we also discuss the features of the cysteine residue modifications and their influence on the efficiency of the search for post-translational modifications and protein products of single nucleotide substitutions.


Assuntos
Artefatos , Cisteína/química , Espectrometria de Massas , Proteômica , Alquilação
2.
J Agric Food Chem ; 68(8): 2506-2515, 2020 Feb 26.
Artigo em Inglês | MEDLINE | ID: mdl-32013414

RESUMO

Thiol groups of cysteine (Cys) residues in proteins react with quinones, oxidation products of polyphenols, to form protein-polyphenol adducts. The aim of the present work was to quantify the amount of adduct formed between Cys residues and 4-methylcatechol (4MC) in minced beef. A Cys-4MC adduct standard was electrochemically synthesized and characterized by liquid chromatography-mass spectrometry (LC-MS) as well as NMR spectroscopy. Cys-4MC adducts were quantified after acidic hydrolysis of myofibrillar protein isolates (MPIs) and LC-MS/MS analysis of meat containing either 500 or 1500 ppm 4MC and stored at 4 °C for 7 days under a nitrogen or oxygen atmosphere. The concentrations of Cys-4MC were found to be 2.2 ± 0.3 nmol/mg MPI and 8.1 ± 0.9 nmol/mg MPI in meat containing 500 and 1500 ppm 4MC, respectively, and stored for 7 days under oxygen. The formation of the Cys-4MC adduct resulted in protein thiol loss, and ca. 62% of the thiol loss was estimated to account for the formation of the Cys-4MC adduct for meat containing 1500 ppm 4MC. Furthermore, protein polymerization increased in samples containing 4MC as evaluated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE), and the polymerization was found to originate from protein-polyphenol interactions as evaluated by a blotting assay with staining by nitroblue tetrazolium.


Assuntos
Cisteína/química , Guaiacol/química , Carne/análise , Fenol/química , Animais , Bovinos , Proteínas Musculares/química , Oxirredução , Quinonas/química , Espectrometria de Massas em Tandem
3.
Food Chem ; 312: 126122, 2020 May 15.
Artigo em Inglês | MEDLINE | ID: mdl-31901825

RESUMO

This study investigated how l-lysine/l-arginine/l-cysteine improved cured sausage color. Visible detection showed that Mb(Fe2+)NO did not form when MetMb(Fe3+) was treated with a combination of l-lysine and NaNO2, l-arginine and NaNO2, or l-cysteine and NaNO2. Visible spectra and electron paramagnetic resonance (EPR) detection together indicated formation of Mb(Fe2+)O2 when MetMb(Fe3+) was treated with l-lysine, l-arginine, or l-cysteine; Mb(Fe2+)NO was formed when MetMb(Fe3+) was treated with a combination of l-lysine and NO, l-arginine and NO, or l-cysteine and NO. Visible, EPR, and fourier transform infrared results suggested formation of a complex of Mb(Fe2+) and l-cysteine by the coordination of sulfydryl and ferrous porphyrin. Therefore, l-lysine, l-arginine, or l-cysteine can promote the formation of Mb(Fe2+)NO by reducing MetMb(Fe3+) to Mb(Fe2+)O2, and l-cysteine promotes formation of a complex of Mb(Fe2+) and l-cysteine via coordination of sulfydryl and ferrous porphyrin, which may improve cured sausage color.


Assuntos
Produtos da Carne/análise , Arginina/química , Cor , Cisteína/química , Espectroscopia de Ressonância de Spin Eletrônica , Lisina/química
4.
Chemistry ; 26(7): 1535-1547, 2020 Feb 03.
Artigo em Inglês | MEDLINE | ID: mdl-31663171

RESUMO

The zinc finger protein tristetraprolin (TTP) regulates inflammation by downregulating cytokine mRNAs. Misregulation results in arthritis, sepsis and cancer, and there is an interest in modulating TTP activity with exogenous agents. Gold has anti-inflammatory properties and has recently been shown to modulate the signaling pathway that produces TTP, suggesting that TTP may be a target of gold. The reactivity of [AuIII (terpy)Cl]Cl2 with TTP was investigated by UV/Vis spectroscopy, spin-filter inductively coupled plasma mass spectrometry, X-ray absorption spectroscopy and native electrospray ionization mass spectrometry. AuIII was found to replace zinc in the protein active site in the reduced AuI form, with the AuI ion coordinated to two cysteine residues in a linear geometry. The replacement of ZnII with AuI results in loss of both secondary structure and RNA binding function. In contrast, when ZnII TTP is bound to its RNA target, no replacement of ZnII with AuI is observed, even in the presence of excess AuIII terpy. This discovery of differential reactivity of gold with TTP versus TTP/RNA offers a potential strategy for selective targeting with gold complexes to control inflammation.


Assuntos
Cisteína/química , Citocinas/química , RNA Mensageiro/metabolismo , RNA/química , Tristetraprolina/química , Humanos , Inflamação , Compostos Organoáuricos/química , RNA Mensageiro/química , RNA Mensageiro/genética , Tristetraprolina/genética , Tristetraprolina/metabolismo , Dedos de Zinco
5.
J Photochem Photobiol B ; 202: 111669, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-31739258

RESUMO

Herein we report the synthesis and characterization of the antifouling Gadolinium oxide (Gd2O3) nanoparticles (NPs) modified with PEG with improved biocompatibility for MR imaging purposes. In this report, using the solvothermal decomposition of Gadolinium (III) in the presence of Na3cit, monitored by surface modification with PEG and L-Cys. The synthesized nanoparticles were confirmed by the TEM, DLS and UV-Visible spectroscopy. The morphological results show normal distance across of the flawless Gd2O3-PEG-Cys-NPs show 7.9 ±â€¯0.4 nm, discretely, with a thin size exchange. This infers the surface adjustment does not obviously alteration the center size of the Gd2O3-NPs when contrasted with the perfect sodium citrate-balanced out Gd2O3-NPs. The Gd2O3-PEG-L-Cys-NPs are highly stable at room temperature, water dispersible and importantly less cytotoxic at high concentration of the NPs. The T1-weighted MR phantasm readings evidentially displayed that the formed PEG coated Gd2O3-PEG and Gd2O3-PEG-Cys-NPs with and without Cys may be performed as the promising T1-weighted MR imaging. The NPs displays no signs of toxicity against the human blood, which represents the biocompatibility for the human medicine applications. The Gd2O3-PEG-Cys-NPs shows relatively, high r1 acceptable cytocompatibility, target specific cancer cells and activate the dual mode MR imaging of lung metastasis cancer model in vitro. The development of versatile zwitterion functionalized Gd2O3 may be promising as an active nanoparticle probe for improved multi-model of MR imaging agents for various cancer diseases.


Assuntos
Meios de Contraste/química , Gadolínio/química , Neoplasias Pulmonares/diagnóstico por imagem , Imagem por Ressonância Magnética , Nanopartículas/química , Polietilenoglicóis/química , Animais , Materiais Biocompatíveis/química , Materiais Biocompatíveis/farmacologia , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Cisteína/química , Hemólise/efeitos dos fármacos , Humanos , Neoplasias Pulmonares/secundário , Melanoma Experimental/patologia , Camundongos , Nanopartículas/toxicidade , Células RAW 264.7 , Transplante Homólogo
6.
Biosci Biotechnol Biochem ; 84(3): 445-454, 2020 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-31771431

RESUMO

Galactose oxidase and amine oxidase contain a cofactor which is generated by post-translational chemical modification to the corresponding amino acid side chains near the copper active center. Such cofactors provide proteins unusual catalytic ability that canonical amino acids cannot exert as well as their structural stability, and thereby are called as protein-derived cofactors. These cofactors and modifications are mostly derived from aromatic amino acid residues, especially Tyr, Trp, and His. Current information about unusual cofactors derived from two of those, heteroaromatic residues (Trp and His) is summarized, especially chemical properties and maturation process of the cross-links between cysteine and heteroaromatic amino acids (His-Cys and Trp-Cys cross-links).Abbreviations: FMN: flavin mononucleotide; FAD: flavin adenine nucleotide; RNA: ribonucleic acid; PDC: protein-derived cofactor; GFP: green fluorescent protein; MIO: 3,5-dihydro-5-methylidene-4-imidazol-4-one; LTQ: lysyl tyrosylquinone; CTQ: cysteine tryptophylquinone; TTQ: tryptophan tryptophylquinone; E.coli: Escherichia coli; WT: wild type.


Assuntos
Reagentes para Ligações Cruzadas/química , Cisteína/química , Histidina/química , Processamento de Proteína Pós-Traducional , Triptofano/química
7.
Chem Commun (Camb) ; 56(1): 74-77, 2020 Jan 04.
Artigo em Inglês | MEDLINE | ID: mdl-31790117

RESUMO

We developed a new method for the de novo formation of fluorophores based on citrate (DNFC) in biological samples. Use of an amide coupling reagent and microwave irradiation greatly facilitates the fluorophore formation on peptides and proteins with N-terminal cysteine or serine. Since N-terminal cysteine and serine can form thiazolopyridone- or oxazolopyridone-based fluorophores emitting blue and green fluorescence, respectively, by the DNFC staining, each organelle, cell and tissue exhibited a characteristic fluorescence distribution. The DNFC staining is able to provide a new potential protocol for future cell imaging, histology and diagnosis.


Assuntos
Corantes Fluorescentes/metabolismo , Sondas Moleculares/metabolismo , Peptídeos/metabolismo , Proteínas/metabolismo , Animais , Linhagem Celular Tumoral , Ácido Cítrico/metabolismo , Cisteína/química , Fluorescência , Corantes Fluorescentes/química , Células HEK293 , Humanos , Camundongos , Microscopia Confocal/métodos , Microscopia de Fluorescência/métodos , Sondas Moleculares/química , Células NIH 3T3 , Peptídeos/química , Estudo de Prova de Conceito , Proteínas/química , Piridonas/química , Piridonas/metabolismo , Serina/química , Tiazóis/química , Tiazóis/metabolismo
8.
J Agric Food Chem ; 68(1): 250-257, 2020 Jan 08.
Artigo em Inglês | MEDLINE | ID: mdl-31823602

RESUMO

Cysteine is a commercially important sulfur-containing amino acid widely used as a supplement in the agricultural and food industries. It is extremely desirable to achieve a high sulfur conversion rate in the fermentation-based cysteine production. Here, the metabolic engineering of Escherichia coli was performed to enhance the sulfur conversion rate in cysteine biosynthesis. Accordingly, the reduction of sulfur loss by the regulator decR and its yhaOM operons were deleted. serACB was integrated into chromosome with constitutive promoter to coordinately increase sulfur utilization. The sulfur assimilation pathways and sulfur transcriptional regulator cysB were overexpressed to regulate sulfur metabolism and enhance sulfur conversion significantly. After the process optimization in fed-batch fermentation, LH16 [SLH02 ΔyhaM Ptrc1-serACB-cysM-nrdH-(pLH03, pTrc99a-cysB)] produced 7.5 g/L of cysteine with a sulfur conversion rate of 90.11%. These results indicate that cysteine production by LH16 is a valuable process in the agricultural and food industries.


Assuntos
Cisteína/biossíntese , Escherichia coli/genética , Escherichia coli/metabolismo , Enxofre/metabolismo , Cisteína/química , Escherichia coli/química , Fermentação , Cinética , Engenharia Metabólica , Óperon , Enxofre/química
9.
Biosens Bioelectron ; 151: 111970, 2020 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-31868609

RESUMO

Herein, a credible construction strategy to improve electrochemiluminescence (ECL) of luminol was developed based on Cu2O-Au heterostructures. Summarily, gold nanoparticles (AuNPs) were anchored on surface of Cu2O nanocube (Cu2O@AuNPs) by spontaneous reduction reaction. Then, luminol molecules were concentrated on Cu2O@AuNPs using L-Cysteine (Cys) as covalent linkage to build the composite emitter (Cu2O@AuNPs-Cys-luminol). The enhancement mechanism was realized by following aspects: (I) Cu2O@AuNPs worked as electrocatalyst for glucose to generate coreactant of H2O2 in situ, avoiding the instability of direct addition of H2O2. (II) luminol molecules were firmly attached on Cu2O@AuNPs to achieve centralized and strong luminescence at low consumption. (III) Cys acted as an intramolecular coreactant and directly linked to luminol to increase luminous efficiency. To validate the effectiveness, a sandwiched immunoassay was built using concanavalinA (ConA) as analyte. Electroreduced graphene film as substrate provided phenoxy-derivatized dextran (DexP) with abundant binding sites and improved conductivity. To improve the specificity, DexP was used to identify ConA via the specific carbohydrate-ConA interaction. Then, Cu2O@AuNPs-Cys-luminol was modified on electrode as ECL signal indicator. The ECL immunosensor achieved determination of ConA with low detection limit of 2.9 × 10-5 ng/mL and excellent stability of continuous potential scan for 8 cycles. Experimental results demonstrated that the proposed construction strategy made considerable progress in ECL efficiency and stability of luminol. The creational pattern of construction strategy achieves high detection capabilities to ConA and expands the applicability of luminol in ECL system. It is expected to have more potential application value in immunoassay with universality.


Assuntos
Cobre/química , Ouro/química , Luminol/química , Nanopartículas Metálicas/química , Técnicas Biossensoriais , Concanavalina A/análise , Cisteína/química , Dextranos/química , Técnicas Eletroquímicas , Eletrodos , Peróxido de Hidrogênio/química , Limite de Detecção , Medições Luminescentes , Oxirredução , Sensibilidade e Especificidade , Propriedades de Superfície
10.
Food Chem ; 307: 125554, 2020 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-31648176

RESUMO

The reaction of Nε-(carboxymethyl) lysine (CML) with eight kinds of non-flavonoid o-benzoquinones and five kinds of flavonoid o-benzoquinones were investigated by cyclic voltammetry at pH 5.0, 7.0 and 8.0 and scan rate of 10, 50 and 100 mV/s. The reactivity of o-benzoquinones towards CML is weakened by the electron-donating substituent and strengthened by the electron-withdrawing substituent on the o-benzoquinone rings. The steric hindrance of the substituents on o-benzoquinone rings also weakens the quinone reactivity. Reaction of 4-methylbenzoquinone with CML (38.0 ±â€¯1.3%) was found to be faster than that with l-lysine (31.3 ±â€¯1.5%) and Nα-acetyl-l-lysine (14.5 ±â€¯0.1%) but slower than that with l-cysteine (≥100.0%) and Nα-acetyl-l-cysteine (≥100.0%) at pH 7.0 and scan rate of 10 mV/s. Products obtained by the reaction of CML with o-benzoquinones were found to include a CML-quinone adduct according to the cyclic voltammetry and UPLC-QTOF-MS/MS analysis.


Assuntos
Benzoquinonas/química , Lisina/análogos & derivados , Catecóis , Cisteína/química , Flavonoides , Lisina/química
11.
J Pharm Biomed Anal ; 177: 112841, 2020 Jan 05.
Artigo em Inglês | MEDLINE | ID: mdl-31522095

RESUMO

For years, d-amino acids were thought to have a minor function in biological processes compared to that of l-enantiomers. Recently, many studies have shown that d-amino acids are present in high concentrations in microorganisms, plants, mammals and humans and execute specific biological functions. One relevant example is that of d-cysteine, whose hydrogen sulfide-producing properties have been found to protect neurons against oxidative stress and to promote dendritic development. Herein, we introduce a chiral LCMS method for the rapid determination of cysteine enantiomers under polar ionic elution conditions (MeOH/MeCN/H2O 49/49/2 v/v/v, containing 50 mM formic acid and 50 mM ammonium formate) developed on a Chiralpak® ZWIX(+) chiral stationary phase. Cysteine enantiomers were analysed in biological samples after efficient reduction of the disulfide bond in cystine; the latter was achieved with the use of 1,4-dithio-dl-threitol as a reducing agent. A baseline resolution (RS = 2.7) was obtained, and the d-enantiomer eluted before the l-enantiomer. For the enantioselective analysis, cysteine was labelled with AccQ-Tag reagent, resulting in improved chromatographic behaviour and MS detection sensitivity. The method was validated according to the Food and Drug Administration guidelines. Good linearity was determined in the ranges of 0.05-0.50 mg/L for d-cysteine and 0.11-0.56 mg/L for l-cysteine. The repeatability and intermediate precision were found to be lower than 4.0%, with trueness ranging from 95.6 to 100.2% for both enantiomers. The LOD and LOQ values were 0.02 and 0.05 mg/L for d-cysteine and 0.04 and 0.11 mg/L for l-cysteine, respectively. The method was successfully applied to cell culture samples treated with d-cysteine.


Assuntos
Cisteína/análise , Espectrometria de Massas/métodos , Células A549 , Técnicas de Cultura de Células , Cromatografia Líquida de Alta Pressão/métodos , Cisteína/química , Humanos , Limite de Detecção , Oxirredução , Reprodutibilidade dos Testes , Estereoisomerismo
12.
Chem Commun (Camb) ; 56(6): 956-959, 2020 Jan 21.
Artigo em Inglês | MEDLINE | ID: mdl-31858094

RESUMO

On-resin intramolecular native chemical ligation (NCL) assisted by N-ethylcysteine using Fmoc/SPPS to obtain cyclic peptides is described. N-terminal cysteine-containing peptides were subjected to NCL conditions leading to cyclization-cleavage reactions and consecutive S → N shift, rendering cyclic peptides in good yields and purities. The compounds were evaluated against P. falciparum 3D7.


Assuntos
Peptídeos Cíclicos/síntese química , Compostos de Sulfidrila/química , Cisteína/análogos & derivados , Cisteína/química , Estrutura Molecular , Peptídeos Cíclicos/química , Resinas Sintéticas/química
13.
Talanta ; 206: 120177, 2020 Jan 01.
Artigo em Inglês | MEDLINE | ID: mdl-31514882

RESUMO

Two highly selective OFF-ON isomer fluorescent probes (1 and 2) for homo-/cysteine were designed and synthesized. The pyrene modified tetraphenylethylene derivative with AIE was used as luminescent group while maleimide was used as recognition group. These two isomer probes were found to be nearly nonfluorescent when treated with GSH. However, upon interaction with Cys or Hcy, the fluorescence was enhanced by 2000 folds in a wide pH range from 3 to 10. Experimental results and DFT calculation have demonstrated that the fluorescence OFF-ON switch of such thiol probes is resulted from the termination of the PET (photo-induced electron transfer) effect through the Michael addition reaction of maleimide unit and thiols. In addition, probe 1 and 2 exhibit excellent selectivity and sensitivity towards Cys, Hcy over GSH and other amino acids, which was confirmed by mass MS. We suggested that Michael addition reaction of these probes with GSH was prevented because of the stereo-hindrance effect. Furthermore, these two isomer probes were successfully used for imaging biothiols in living H1299 lung cancer cells.


Assuntos
Cisteína/análise , Corantes Fluorescentes/química , Glutationa/química , Homocisteína/análise , Linhagem Celular Tumoral , Cisteína/química , Teoria da Densidade Funcional , Fluorescência , Corantes Fluorescentes/síntese química , Homocisteína/química , Humanos , Maleimidas/síntese química , Maleimidas/química , Microscopia de Fluorescência/métodos , Modelos Químicos , Estilbenos/síntese química , Estilbenos/química
14.
Phytochemistry ; 169: 112188, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-31683228

RESUMO

The metabolism of the phytoalexin rapalexin A, a unique indole isothiocyanate (ITC) produced by crucifers (family Brassicaceae), was investigated. Three phytopathogenic fungal species were examined: Colletotrichum dematium (Pers.:Fr.) Grove, a broad host range pathogen, C. higginsianum Sacc., a host-selective pathogen of crucifers and C. lentis Damm, a host-selective pathogen of lentils (Lens culinaris Medik.). The metabolism of rapalexin A by C. dematium and C. higginsianum was similar, taking place via one common intermediate and two divergent pathways, but C. lentis was unable to transform rapalexin A. Both C. higginsianum and C. dematium transformed rapalexin A to two previously undescribed metabolites, the structures of which were confirmed by chemical synthesis: N-acetyl-S-(8-methoxy-4H-thiazolo[5,4-b]indol-2-yl)-L-cysteine and 4-hydroxy-3-(4-methoxy-1H-indol-3-yl)-2-thioxothiazolidine-4-carboxylic acid. That is, both fungal pathogens metabolized and detoxified rapalexin A by addition of the thiol group of L-Cys residue to the isothiocyanate carbon of rapalexin A, a transformation usually catalyzed by glutathione transferases. Coincidentally, this metabolic pathway is employed by mammals and insects to detoxify isothiocyanates and other xenobiotics. Hence, C. higginsianum could be a useful model fungus to uncover genes involved in the detoxification pathways of ITCs and related xenobiotics. Our overall results suggest that increasing rapalexin A production in specific crucifers could increase crop resistance to certain fungal pathogens.


Assuntos
Colletotrichum/metabolismo , Cisteína/metabolismo , Isotiocianatos/metabolismo , Sesquiterpenos/metabolismo , Acetilação , Brassicaceae/química , Brassicaceae/metabolismo , Ciclização , Cisteína/química , Isotiocianatos/química , Estrutura Molecular , Estresse Oxidativo , Sesquiterpenos/química
15.
Food Chem ; 309: 125697, 2020 Mar 30.
Artigo em Inglês | MEDLINE | ID: mdl-31727513

RESUMO

Formation of green trihydroxy benzacridine (TBA) derivatives when chlorogenic acid (CGA) quinones and amino acids react can be unappealing for some consumers. Cysteine was studied as an anti-greening strategy, given that cysteine-CGA conjugates are colorless. Buffered 2.55 mM CGA: 5.09 mM lysine: (0-5.09) mM cysteine solutions at pH 8 and 9 were prepared and incubated for a maximum of 48 h at 22 C. Color intensity and conjugate formation was monitored spectrophotometrically, and by HPLC and LC-MS respectively, while antioxidant capacity was measured by Folin-Ciolcateau and Trolox equivalent antioxidant capacity assays. Green TBA formation was promoted at higher pH and inhibited as cysteine concentration increased. Concentration-dependent cysteine inhibition of CGA-lysine greening was attributed to redox diphenol regeneration and formation of cysteinyl-CGA conjugates, which also contributed to antioxidant capacity. pH had a greater effect on antioxidant capacity than added cysteine. Results suggested a potential anti-greening approach for alkaline CGA quinone-amine greening.


Assuntos
Antioxidantes/química , Benzoquinonas/química , Cisteína/química , Ácido Clorogênico/química , Cromatografia Líquida de Alta Pressão , Cor , Reação de Cicloadição , Concentração de Íons de Hidrogênio , Espectrometria de Massas , Oxirredução
16.
Anal Bioanal Chem ; 412(4): 833-840, 2020 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-31872274

RESUMO

Accurate determination of the drug-to-antibody ratio (DAR) of interchain cysteine-linked antibody-drug conjugates (ADCs) is challenging. High-resolution mass spectrometry (HRMS) analysis of the ADCs at the intact or subunit level provides a feasible way to measure the DAR. However, the measured DAR is usually lower than the true DAR because of the variation in ionization efficiency between different DAR species. In this work, we developed a novel standard-free HRMS method involving isotope-labeled payload conjugation, protease digestion, and liquid chromatography-HRMS (LC-HRMS) analysis for accurate determination of the DAR of the interchain cysteine-linked ADCs with cleavable or non-cleavable linkers. Isotope-labeled payload conjugations eliminated the structural and chemical differences between different DAR species and ensured that the drugs or payload-containing peptides could be separated from each other in the mass spectrometer. A papain digestion strategy for ADCs with cleavable linkers showed a DAR of 3.79, with a relative standard deviation (RSD) of 0.48 (n = 3). Similarly, the trypsin and chymotrypsin digestion strategy that is applicable to ADCs with non-cleavable linkers showed a DAR of 3.77 and an RSD of 0.86 (n = 3). The DAR determined by this method was consistent with the DAR of the ADCs that was measured by the UV/Vis method. This method will be very useful to researchers working in the field of ADC discovery and development. Graphical abstract.


Assuntos
Cisteína/química , Imunoconjugados/química , Cromatografia Líquida/métodos , Espectrometria de Massas/métodos , Subunidades Proteicas/química , Proteólise
17.
Analyst ; 144(24): 7242-7249, 2019 Dec 21.
Artigo em Inglês | MEDLINE | ID: mdl-31687669

RESUMO

Rapid detection and identification of bacteria is important for human health, biodefense, and food safety. Small arrays of different antimicrobial peptides (AMPs) enable the identification of lipopolysaccharide (LPS) samples from a variety of bacterial species and strains. A model system for examining how peptide presentation affects LPS detection is the sheep myeloid antimicrobial peptide (SMAP-29), which contains a helix-turn-helix motif. Varying the cysteine attachment site on SMAP-29 controls the three-dimensional presentation of the peptide on the surface, altering the ability of the peptide to discriminate between LPS samples. A small array of only SMAP-29 variants-and no other peptides-is capable of discriminating among LPS samples from multiple bacterial species, as well as between different strains within the same species, with high accuracy.


Assuntos
Peptídeos Catiônicos Antimicrobianos/química , Proteínas Sanguíneas/química , Catelicidinas/química , Lipopolissacarídeos/química , Animais , Peptídeos Catiônicos Antimicrobianos/farmacologia , Bactérias/metabolismo , Sítios de Ligação , Cisteína/química , Humanos , Lipopolissacarídeos/antagonistas & inibidores , Ovinos , Especificidade da Espécie , Ressonância de Plasmônio de Superfície
18.
J Food Sci ; 84(12): 3584-3593, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31721210

RESUMO

Maillard reaction intermediate (MGX) generated from glutathione and xylose in aqueous medium was prepared via the Maillard reaction performed under a two-stage temperature increase process. The purified MGX was identified by Fourier-transform infrared spectroscopy, mass spectrometry, and nuclear magnetic resonance as N-(1-deoxy-d-xylulos-1-yl)-glutathione (Amadori compound, C15 H25 O10 N3 S) with five main isomers. The method of Maillard reaction performed under a two-stage temperature increase process was further verified by high-performance liquid chromatography. The optimal reaction time and temperature for the preparation of MGX was determined as 60 min at 90 °C. The yield of MGX was increased from 8.60% to 55.52% through thermal reaction coupled with vacuum dehydration. In addition, rapid and more Maillard-type volatile compounds were formed in MGX during thermal treatment than that in Maillard reaction products or glutathione-xylose mixture. Beside, MGX possessed more pleasing meat-like volatile profile compared with the Amadori compound of glutamic acid-xylose (AAX), cysteine-xylose (ACX), and glycine-xylose (AGX). Therefore, it suggested that the MGX had the potential to achieve a better flavor formation during thermal treatment. PRACTICAL APPLICATION: Maillard reaction intermediates, such as Amadori or Heyns rearrangement products (ARP or HRP), are important flavor precursors, which possess stable physicochemical properties, but tend to degrade into flavor compounds at high temperatures. Maillard reaction intermediate from glutathione and xylose acts as primary flavor enhancers to complete Maillard reaction to produce flavors in the subsequent thermal processing, which can significantly improve and stabilize the flavor quality of the meaty food, and deserves a very broad application prospects. The new developed method will be a significant theoretical basis on research preparation and properties of Maillard reaction intermediates in complex food systems.


Assuntos
Glutationa/química , Xilose/química , Cisteína/química , Aromatizantes/química , Ácido Glutâmico/química , Produtos Finais de Glicação Avançada/química , Glicina/química , Temperatura Alta , Reação de Maillard , Vácuo
19.
Chem Commun (Camb) ; 55(91): 13725-13728, 2019 Nov 12.
Artigo em Inglês | MEDLINE | ID: mdl-31660558

RESUMO

In this communication, the mechanism of how surface chirality affects amyloid-ß (Aß) fibrillation was firstly unravelled at the molecular level: a chiral surface serves to control the 2D-diffusion and surface residence time of Aß molecules via the chiral recognition with Aß to allow precursor Aß to laterally diffuse and collide with each other for oligomerization and fibrillation. Surface chirality that shortens the surface residence time of Aß, for example, R-cysteine modification with carboxylic, secondary amine and thiol groups surrounding the chiral center, can retard Aß oligomerization and fibrillation. This work is essential to a deeper fundamental understanding of the effects of surface chirality on amyloidosis processes as well as the development of chiral materials to inhibit Aß fibrillation.


Assuntos
Peptídeos beta-Amiloides/metabolismo , Amiloide/metabolismo , Fragmentos de Peptídeos/metabolismo , Amiloide/química , Peptídeos beta-Amiloides/química , Cisteína/química , Ligações de Hidrogênio , Microscopia de Força Atômica , Fragmentos de Peptídeos/química , Dióxido de Silício/química , Estereoisomerismo
20.
Molecules ; 24(19)2019 Oct 02.
Artigo em Inglês | MEDLINE | ID: mdl-31581645

RESUMO

In recent years, several catalyst-free site-specific reactions have been investigated for the efficient conjugation of biomolecules, nanomaterials, and living cells. Representative functional group pairs for these reactions include the following: (1) azide and cyclooctyne for strain-promoted cycloaddition reaction, (2) tetrazine and trans-alkene for inverse-electron-demand-Diels-Alder reaction, and (3) electrophilic heterocycles and cysteine for rapid condensation/addition reaction. Due to their excellent specificities and high reaction rates, these conjugation methods have been utilized for the labeling of radioisotopes (e.g., radiohalogens, radiometals) to various target molecules. The radiolabeled products prepared by these methods have been applied to preclinical research, such as in vivo molecular imaging, pharmacokinetic studies, and radiation therapy of cancer cells. In this review, we explain the basics of these chemical reactions and introduce their recent applications in the field of radiopharmacy and chemical biology. In addition, we discuss the significance, current challenges, and prospects of using bioorthogonal conjugation reactions.


Assuntos
Antineoplásicos/síntese química , Compostos Radiofarmacêuticos/síntese química , Animais , Antineoplásicos/química , Catálise , Química Click , Reação de Cicloadição , Cisteína/química , Humanos , Imagem Molecular , Estrutura Molecular , Neoplasias/tratamento farmacológico , Neoplasias/metabolismo , Compostos Radiofarmacêuticos/química
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