NMR Verification of Structures of Bacterial Saccharide Precursors for Vaccines

疫苗用细菌糖前体结构的核磁共振验证

• 批准号: 7968700
• 负责人: Rachel Schneerson
• 金额: $ 3.45万
• 依托单位: EUNICE KENNEDY SHRIVER NATIONAL INSTITUTE OF CHILD HEALTH & HUMAN DEVELOPMENT
• 依托单位国家: 美国
• 资助国家: 美国
• 起止时间: 至
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/7968700
• 关键词: Agreement; Antigens; Bacteria; Bacterial Vaccines; Carbon; Chemical Structure; Complex; Coupling; Data; Detection; Excision; Fingerprint; Glucose; Goals; Haemophilus influenzae; Laboratories; Length; Lipopolysaccharides; Literature; Measurement; Measures; Methods; Molecular Structure; Monosaccharides; NMR Spectroscopy; Oligosaccharides; Polysaccharides; Preparation; Proteins; Resolution; Rhamnose; Ribitol; Sampling; Shigella flexneri; Signal Transduction; Structure; Techniques; Vaccines; Validation; Vibrio cholerae; base; interest; mutant; stereochemistry; two-dimensional; vaccine development

A further 20 monosaccharide and oligosaccharide intermediates of Vibrio cholera antigens, for the preparation of a hexasaccharide-protein conjugate have been analyzed by high-resolution one-dimensional (1D) 1H and 1D 13C NMR at 500 MHz and 126 MHz, respectively. Validation of the chemical structures was based on integration of the 1D 1H NMR spectra, and on counting of carbon resonances in the 13C spectra, especially in the well-separated spectral regions of the C=O, aromatic C, C-1, C-N, and C-CH3 moieties. The NMR spectra were also used to assess the purity of the synthetic preparations, for example, determination of the extent of completion of hydrogenolytic removal of benzyl protecting groups by measurement of the content of aromatic groups. NMR spectra measured at 500 MHz have been provided for the identification of six polysaccharide fragments isolated from the lipopolysaccharide of Shigella flexneri type 2a, and seven polysaccharide fractions prepared from a mutant version of this bacterium. Comparison of the integrals of the polysaccharide H-1 and rhamnose methyl signals facilitated detection of core polysaccharide in the samples. The techniques provide complex fingerprint spectra that can be compared with literature data to verify identity. A new preparation of the capsular polysaccharide of Haemophilus influenzae type a has been investigated by 1D 1H and 13C NMR and found to have excellent purity, and close agreement with the spectra of previous preparations in this laboratory. These spectra had been analyzed several years ago by two-dimensional NMR methods to give sets of 1H-1H and C-1,H-1 coupling constants for the D-glucose residue and 13C-31P coupling constants for the D-glucose and D-ribitol components of the polysaccharide. Also obtained, were complete assignments of the 1H and 13C NMR signals of the beta-D-glucopyranosyl and D-ribitol residues, which are joined by a phosphoric diester linkage between O-4 of the D-glucose and O-5 of the D-ribitol.

另外20种单糖和寡糖中间体的中间体，用于制备六糖 - 蛋白质缀合物，分别通过高分辨率的一维（1D）1H和1d 13c NMR分别在500 MHz和126 MHz和126 MHz的情况下进行了分析。化学结构的验证是基于1D 1H NMR光谱的整合，以及在13C光谱中的碳共振计数，尤其是在C = O，芳香族C，C-1，C-N和C-CH3部分的良好分离光谱区域中。 NMR光谱还用于评估合成制剂的纯度，例如确定通过测量芳族基团的含量来确定苄基保护基团的氢解度去除的程度。 已经提供了在500 MHz下测得的NMR光谱，以鉴定从2型志贺氏菌的脂多糖中分离出的六个多糖片段，以及从该细菌的突变版制备的七个多糖级分。多糖H-1和鼠李糖甲基信号的积分的比较促进了样品中核心多糖的检测。这些技术提供了复杂的指纹光谱，可以将其与文献数据进行比较以验证身份。 已经通过1d 1h和13c NMR研究了A型流感嗜血杆菌的囊囊多糖的新制备，发现具有极好的纯度，并且与该实验室先前制剂的光谱密切一致。几年前，通过二维NMR方法对这些光谱进行了分析，以给出D-葡萄糖残基的1H-1H和C-1，H-1偶联常数，以及D-葡萄糖和多糖含量的D-葡萄糖和D-核醇成分的D-葡萄糖残基和13C-31p耦合常数。还获得了β-D-丙糖基和D-核醇残基的1H和13C NMR信号的完整分配，这些信号与D-核糖醇的D-葡萄糖和O-5之间的磷酸二酯连接在一起。