糖生物学--生命科学研究的新热点

糖生物学是糖的化学和生物学研究相结合而产生的一门新兴学科，近20年来，糖生物学研究取得了很多重要成就．介绍了糖生物学的重要进展及热点，包括糖链在新生肽链折叠过程中、在免疫系统中的作用和糖链与微生物感染等．目前许多国家对糖生物学研究都给予了足够的支持和重视，糖生物学已成为生命科学研究的前沿和新热点．     

Conformational analysis of the disaccharide methyl α‐d‐mannopyranosyl‐(1→3)‐2‐O‐acetyl‐β‐d‐mannopyranoside monohydrate

The crystal structure of methyl α‐d ‐mannopyranosyl‐(1→3)‐2‐O‐acetyl‐β‐d ‐mannopyranoside monohydrate, C₁₅H₂₆O₁₂·H₂O, ( II ), has been determined and the structural parameters for its constituent α‐d ‐mannopyranosyl residue compared with those for methyl α‐d ‐mannopyranoside. Mono‐O‐acetylation appears to promote the crystallization of ( II ), inferred from the difficulty in crystallizing methyl α‐d ‐mannopyranosyl‐(1→3)‐β‐d ‐mannopyranoside despite repeated attempts. The conformational properties of the O‐acetyl side chain in ( II ) are similar to those observed in recent studies of peracetylated mannose‐containing oligosaccharides, having a preferred geometry in which the C2—H2 bond eclipses the C=O bond of the acetyl group. The C2—O2 bond in ( II ) elongates by ～0.02 Å upon O‐acetylation. The phi (?) and psi (ψ) torsion angles that dictate the conformation of the internal O‐glycosidic linkage in ( II ) are similar to those determined recently in aqueous solution by NMR spectroscopy for unacetylated ( II ) using the statistical program MA′AT, with a greater disparity found for ψ (Δ = ～16°) than for ? (Δ = ～6°).     

Metabolic Labeling and Imaging of N‐Linked Glycans in Arabidopsis Thaliana

Molecular imaging of glycans has been actively pursued in animal systems for the past decades. However, visualization of plant glycans remains underdeveloped, despite that glycosylation is essential for the life cycle of plants. Metabolic glycan labeling in Arabidopsis thaliana by using N‐azidoacetylglucosamine (GlcNAz) as the chemical reporter is reported. GlcNAz is metabolized through the salvage pathway of N‐acetylglucosamine (GlcNAc) and incorporated into N‐linked glycans, and possibly intracellular O‐GlcNAc. Click‐labeling with fluorescent probes enables visualization of newly synthesized N‐linked glycans. N‐glycosylation in the root tissue was discovered to possess distinct distribution patterns in different developmental zones, suggesting that N‐glycosylation is regulated in a developmental stage‐dependent manner. This work shows the utility of metabolic glycan labeling in elucidating the function of N‐linked glycosylation in plants.     

Total Biosynthesis of Legionaminic Acid,a Bacterial Sialic Acid Analogue

Legionaminic acid, Leg5,7Ac₂, a nonulosonic acid like 5‐acetamido neuraminic acid (Neu5Ac, sialic acid), is found in cell surface glycoconjugates of bacteria including the pathogens Campylobacter jejuni, Acinetobacter baumanii and Legionella pneumophila. The presence of Leg5,7Ac₂ has been correlated with virulence in humans by mechanisms that likely involve subversion of the host's immune system or interactions with host cell surfaces due to its similarity to Neu5Ac. Investigation into its role in bacterial physiology and pathogenicity is limited as there are no effective sources of it. Herein, we construct a de novo Leg5,7Ac₂ biosynthetic pathway by combining multiple metabolic modules from three different microbial sources (Saccharomyces cerevisiae, C. jejuni, and L. pneumophila). Over‐expression of this de novo pathway in Escherichia coli that has been engineered to lack two native catabolic pathways, enables significant quantities of Leg5,7Ac₂ (≈120 mg L^?1 of culture broth) to be produced. Pure Leg5,7Ac₂ could be isolated and converted into CMP‐activated sugar for biochemical applications and a phenyl thioglycoside for chemical synthesis applications. This first total biosynthesis provides an essential source of Leg5,7Ac₂ enabling study of its role in prokaryotic and eukaryotic glycobiology.     

Glycoproteomics: Charting new territory in mass spectrometry and glycobiology

Glycosylation is an incredibly common and diverse post-translational modification that contributes widely to cellular health and disease. Mass spectrometry is the premier technique to study glycoproteins; however, glycoproteomics has lagged behind traditional proteomics due to the challenges associated with studying glycosylation. For instance, glycans dissociate by collision-based fragmentation, thus necessitating electron-based fragmentation for site-localization. The vast glycan heterogeneity leads to lower overall abundance of each glycopeptide, and often, ion suppression is observed. One of the biggest issues facing glycoproteomics is the lack of reliable software for analysis, which necessitates manual validation and serves as a massive bottleneck in data processing. Here, I will discuss each of these challenges and some ways in which the field is attempting to address them, along with perspectives on how I believe we should move forward.     

Metabolic Labeling of Sialic Acids in Living Animals with Alkynyl Sugars

Sialome sweet sialome : As sialic acids are involved in many host–pathogen recognition events and are markers of embryonic and malignant tissues, there is great interest in methods for the enrichment and identification of sialylated glycoproteins from complex tissues. Now N‐(4‐pentynoyl)mannosamine can be used to metabolically label sialylated glycoproteins in living animals, enabling future identification of new biomarkers.

     

糖芯片最新研究进展

糖芯片是一种快速、高效、高通量获取糖-生物大分子相互作用信息的生物检测技术,对后基因组时代糖生物学的发展具有重大影响。本文主要论述了糖芯片固定化技术的最新进展,包括未经化学修饰的糖的化学固定,天然糖库及其固定,复杂寡糖的合成及其固定,糖基的密度差异固定化技术以及间隔基的引入技术。这些新的固定化技术保持了糖的化学结构,扩大了糖芯片的来源和应用范围,进一步提高了糖芯片的检测效率。此外,本文还介绍了虚拟筛选技术在这一领域的应用潜力以及糖芯片在医疗诊断等方面的应用,最后对糖芯片技术遇到的挑战和发展做了展望。     

Resources and Methods for Engineering “Designer” Glycan-Binding Proteins

This review provides information on available methods for engineering glycan-binding proteins (GBP). Glycans are involved in a variety of physiological functions and are found in all domains of life and viruses. Due to their wide range of functions, GBPs have been developed with diagnostic, therapeutic, and biotechnological applications. The development of GBPs has traditionally been hindered by a lack of available glycan targets and sensitive and selective protein scaffolds; however, recent advances in glycobiology have largely overcome these challenges. Here we provide information on how to approach the design of novel “designer” GBPs, starting from the protein scaffold to the mutagenesis methods, selection, and characterization of the GBPs.     

Carbohydrates as the next frontier in pharmaceutical research

Synthetic carbohydrates and glycoconjugates are used to study their roles in biological important processes such as inflammation, cell-cell recognition, immunological response, metastasis, and fertilization. The development of an automated oligosaccharide synthesizer greatly accelerates the assembly of complex, naturally occurring carbohydrates as well as chemically modified oligosaccharide structures and promises to have major impact on the field of glycobiology. Tools such as microarrays, surface plasmon resonance spectroscopy, and fluorescent carbohydrate conjugates to map interactions of carbohydrates in biological systems are presented. Case studies of the successful application of carbohydrates as active agents are discussed, for example, fully synthetic oligosaccharide vaccines to combat tropical diseases (e.g., malaria), bacterial infections (e.g., tuberculosis), viral infections such as HIV, and cancer. Aminoglycosides serve as examples of drugs acting through carbohydrate-nucleic-acid interactions, while heparin works by carbohydrate-protein interactions. A general, modular strategy for the complete stereoselective synthesis of defined heparin oligosaccharides is presented. A carbohydrate-functionalized fluorescent polymer has been shown to detect miniscule amounts of bacteria faster than commonly used methods.     

Involvement of DPY19L3 in Myogenic Differentiation of C2C12 Myoblasts

DPY19L3 has been identified as a C-mannosyltransferase for thrombospondin type-1 repeat domain-containing proteins. In this study, we focused on the role of DPY19L3 in the myogenic differentiation of C2C12 mouse myoblast cells. We carried out DPY19L3 gene depletion using the CRISPR/Cas9 system. The result showed that these DPY19L3-knockout cells could not be induced for differentiation. Moreover, the phosphorylation levels of MEK/ERK and p70S6K were suppressed in the DPY19L3-knockout cells compared with that of parent cells, suggesting that the protein(s) that is(are) DPY19L3-mediated C-mannosylated and regulate(s) MEK/ERK or p70S6K signaling is(are) required for the differentiation.