Quantitative gas chromatography/mass spectrometry determination of C-mannosylation of tryptophan residues in glycoproteins

C-mannosylation of Trp residue is one of the most recently discovered types of glycosylation, but the identification of these mannosylated residues in proteins is rather tedious. In a previous paper, it was reported that the complete analysis of all constituents of glycoproteins (sialic acids, monosaccharides, and amino acids) could be determined on the same sample in three different steps of gas chromatography/mass spectrometry of heptafluorobutyrate derivatives. It was observed that during the acid-catalyzed methanolysis step used for liberation of monosaccharide from classical O- and N-glycans, Trp and His were quantitatively transformed by the addition of a methanol molecule on their indole and imidazole groups, respectively. These derivatives were stable to acid hydrolysis used for the liberation of amino acids. Since monosaccharide derivatives were also stabilized as heptafluorobutyrate derivatives of O-methyl-glycosides, it was suggested that C-mannosides of Trp residues could quantitatively be recovered. Based on the analyses of standard compounds, peptides and RNase 2 from human urine, we report that C((2))-mannosylated Trp could be quantitatively recovered and identified during the step of amino acid analysis. Analyses of different samples indicated that this type of glycosylation is absent in bacteria and yeasts.     

Isolation, fractionation and some properties of polysaccharides produced in a bound form by Azospirillum brasilense and their possible involvement in Azospirillum-wheat root interactions

Abstract It was shown that Azospirillum brasilense strains Sp7, Sp107, Sp245, and S17 when cultivated in a liquid synthetic malate medium to the end of the exponential phase of growth, produced at least two complex polysaccharide-containing components. The components were arbitrarily called lipopolysaccharide-protein complex and polysaccharide-lipid complex. These complexes were shown to interact with a wheat germ agglutinin. From polysaccharide-lipid complexes, acidic polysaccharides were isolated and their specific rotation, molecular masses, affinity for wheat germ agglutinin, and monosaccharide composition were determined. The polysaccharides of all strains contained rhamnose, galacturonic acid, and glucosamine, while the polysaccharides of strains Sp7 and S17 included additional fucose and mannose, respectively, and both had galactose. It is suggested that lipopolysaccharide-protein complexes, polysaccharide-lipid complexes, and polysaccharides may be involved in the process of interaction of azospirilla with wheat root surfaces.     

单糖组成分析方法的研究进展

单糖组成分析是对多糖以及糖蛋白中多种单糖组分的定性、定量分析。该分析方法可被用来分析多糖结构以及在多糖类药物研发或生产中起到质量控制的作用。就近几年来多糖以及糖蛋白中的单糖组成、分析方法进行了综述,包括对样品的水解方法以及不同色谱法和电泳法利弊的分析。     

Development of biofuel cells based on gold nanoparticle decorated multi-walled carbon nanotubes

This study focused on developing the synthesis of Au nanoparticle-decorated functionalized multi-walled carbon nanotubes (Au-NPs/f-MWCNTs) for monosaccharide (bio-fuel) oxidation reactions and practical application in air-biofuel cells. We developed a scalable and straightforward method to synthesize Au-NPs/f-MWCNTs which allow us to control the loading and size of the Au-NPs. The Au-NPs/f-MWCNTs exhibited better catalytic activities and stability than the Au sheet and subsequently resulted in a threefold increase in the power density of the air-glucose fuel cell with an exceptionally high open circuit voltage (∼1.3 V). The catalytic efficiency was confirmed by high performance liquid chromatography with the superior of the Au-NPs/f-MWCNTs over a bare gold electrode. In addition, the application of this advanced catalyst to other monosaccharide oxidation reactions figured out that the configuration of –OH groups at C₂ and C₃ of the reactants plays an important role in the initial adsorption process, and thus, affects the required activation energy for further oxidation. The different monosaccharides lead to significantly different fuel cell performances in terms of power density, which coherently corresponds to the difference in the configuration of C₂ and C₃. Because two small air-glucose fuel cells using Au-NPs/f-MWCNTs can run a LED lamp, further applications of other monosaccharides as fuel in biofuel cells for equivalent required power devices may be possible.     

南瓜多糖的单糖组成及其生理效应

南瓜多糖是南瓜中重要的活性成分,由葡萄糖、半乳糖、阿拉伯糖、葡萄糖醛酸等单糖组成,是一类主链上含有β-1,3糖苷键、侧链含有α-糖苷键的杂多糖。南瓜多糖具有降血糖、调理血脂、抑制肿瘤、抗氧化等生理作用。本文对南瓜多糖的化学组成和生理效应进行了综述,最后,对南瓜多糖的研究方向进行了展望。     

Synthesis and utility of sulfated chromogenic carbohydrate model substrates for measuring activities of mucin-desulfating enzymes

A chromogenic substrate, 4-nitrophenyl 2-acetamido-2-deoxy-beta-D-glucopyranoside 6-sodium sulfate was synthesized and used in combination with beta-N-acetylhexosaminidase for detection of the sulfatase, MdsA, by release of 4-nitrophenol. MdsA was originally isolated from the bacterium Prevotella strain RS2 and is believed to be involved in desulfation of sulfomucins, major components of the mucus barrier protecting the human colon surface. The exo nature of the MdsA sulfatase was indicated by its inability to de-esterify the disaccharide 4-nitrophenyl beta-D-galactopyranosyl-(1-->4)-2-acetamido-2-deoxy-beta-D-glucopyranoside 6-sodium sulfate. This latter compound was prepared from monosaccharide precursors by two different methods, the shorter requiring just six steps from 4-nitrophenyl 2-acetamido-2-deoxy-beta-D-glucopyranoside and giving an overall yield of 26.4%. The syntheses of 4-nitrophenyl beta-D-galactopyranoside 3-triethylammonium sulfate and 6-triethylammonium sulfate and their use in combination with beta-galactosidase as chromogenic substrates for detecting Bacteroides fragilis sulfatases with different specificities was also demonstrated.     

黄精多糖组成及其抗氧化活性分析

分离纯化湖南新化产黄精多糖,并对其组分、结构特征和抗氧化活性进行研究。本研究采用碱提法、Sevag法脱蛋白得水溶性粗多糖(PSP),并经DEAE-52和Sepharose CL-6B色谱柱分离纯化得黄精多糖分级组分。通过紫外、红外光谱和GC-MS进行初步结构分析,并检测其体内外抗氧化能力。结果 PSP经纯化得1个组分PSP-1a,不含蛋白质和核酸,具有典型多糖吸收峰,主要由半乳糖、阿拉伯糖、鼠李糖、木糖、葡萄糖组成,其摩尔百分含量分别为0.18%、1.50%、97.25%、0.77%和0.3%。体外抗氧化活性研究表明:PSP和PSP-1a均具有一定抗氧化性能力,且随浓度(1~10 mg/L)增大而增强,PSP-1a的抗氧化能力优于PSP,但弱于VC。体内实验显示,PSP-1a改善了高脂膳食肥胖小鼠结肠匀浆组织的氧化应激状况。     

Total synthesis of phenylpropanoid glycosides, grayanoside A and syringalide B, through a common intermediate

Phenylpropanoid glycosides are known as bioactive natural products. Two of them, grayanoside A (1) and syringalide B (2), were synthesized through a common intermediate, using benzyl as temporary protecting group following a shorter route.     

Molecular cloning and expression analysis of a monosaccharide transporter gene OsMST4 from rice (Oryza sativa L.)

Two putative glycosyltransferases in Arabidopsis thaliana, designated reduced residual arabinose-1 and -2 (RRA1 and RRA2), are characterized at the molecular level. Both genes are classified in CAZy GT-family-77 and are phylogenetically related to putative glycosyltranferases of Chlamydomonas reinhardtii. The expression pattern of the two genes was analyzed by semi-quantitative RT-PCR using mRNA extracted from various organs of bolting Arabidopsis thaliana plants. In addition, promoter::gusA analysis of transgenic Arabidopsis thaliana containing a fusion between either the RRA-1 or -2 promoter fragment and the gusA reporter gene showed that whereas the RRA1 promoter was primarily active in the apical meristem, the expression pattern of the RRA2 promoter was more diverse but also highly active in the meristematic region. In addition, T-DNA mutant insertion lines of both RRA-1 and -2, were identified and characterized at the molecular and biochemical level. Monosaccharide compositional analyses of cell wall material isolated from the meristematic region showed a ca. 20% reduction in the arabinose content in the insoluble/undigested cell wall residue after enzymatic removal of xyloglucan and pectic polysaccharides. These data indicate that both RRA-1 and -2 play a role in the arabinosylation of cell wall component(s).