Structural elucidation of glycosaminoglycans through characterization of disaccharides obtained after fragmentation by hydrazine-nitrous acid treatment.
Structural elucidation of glycosaminoglycans through characterization of disaccharides obtained after fragmentation by hydrazine-nitrous acid treatment.
复制标题
通过对肼-亚硝酸处理断裂后获得的二糖进行表征，阐明糖胺聚糖的结构。
DOI：
10.1016/0003-9861(85)90063-3
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发表时间：
1985
影响因子：
3.9
通讯作者：
Spiro,RG
中科院分区：
文献类型：
--
作者：
Edge,AS;Spiro,RG
肼解糖胺聚糖，使N-脱乙酰基，然后用亚硝酸处理，在交替的己糖胺残基上进行脱氨基裂解，已被用于鉴定和定量二糖序列和这些聚合物中纳克量的O-硫酸酯取代位置。在通过NaB 3 H4还原进行放射性标记后，肼-亚硝酸产物在Dowex 1上分级分离，并通过薄层色谱法进一步分离为二糖，以硫酸化或未硫酸化的脱水甘露醇或脱水塔罗糖醇终止。透明质酸、硫酸角质素、4-硫酸软骨素、6-硫酸软骨素、硫酸皮肤素和肝素的片段化产生总共14种二糖，其包含哺乳动物糖胺聚糖中存在的主要序列（大于1摩尔%）。代表硫酸软骨素[GlcUAβ1 → 3脱水Talitol（4-SO 4）和GlcUAβ1 → 3脱水Talitol（6-SO 4）]以及硫酸皮肤素链[IdUAα1 → 3脱水Talitol（4-SO 4）和GlcUAβ1 → 3脱水Talitol（4-SO 4）]链的主要变体的二糖可通过该方法轻松定量。在肝素的情况下，通过直接亚硝酸和肼-亚硝酸处理产生的二糖的比较还提供了N-硫酸基团分布的评估。史密斯高碘酸降解和糖苷酶digestriol的各种二糖的表征促进了制备和薄层色谱分辨率的完整系列的单硫酸衍生物的脱水甘露醇和脱水talitol;硫酸酯被证明是稳定的肼和亚硝酸处理。肼-亚硝酸片段化程序的高灵敏度应证明可用于细胞表面和基底膜蛋白聚糖以及少量存在的其他硫酸化糖缀合物的结构阐明。
Hydrazinolysis of glycosaminoglycans to bring aboutN-deacetylation followed by nitrous acid treatment to effect deaminative cleavage at alternating hexosamine residues has been used to make possible identification and quantitation of disaccharide sequences and position ofO-sulfate substitution in nanogram amounts of these polymers. After radiolabeling by NaB3H4reduction the hydrazine-nitrous acid products were fractionated on Dowex 1 and further resolved by thin-layer chromatography into disaccharides terminating in either sulfated or unsulfated anhydromannitol or anhydrotalitol. Fragmentation of hyaluronic acid, keratan sulfate, chondroitin 4-sulfate, chondroitin 6-sulfate, dermatan sulfate, and heparin yielded a total of 14 disaccharides comprising the major sequences (greater than 1 mol%) occurring in mammalian glycosaminoglycans. Disaccharides representing the predominant variants of the chondroitin sulfates [GlcUAβ1 → 3anhydrotalitol(4-SO4) and GlcUAβ1 → 3anhydrotalitol(6-SO4)] as well as of dermatan sulfate chains [IdUAα1 → 3anhydrotalitol(4-SO4) and GlcUAβ1 → 3anhydrotalitol(4-SO4)] chains could readily be quantitated by this approach. In the case of heparin a comparison of the disaccharides produced by direct nitrous acid and hydrazine-nitrous acid treatments moreover provided an assessment of the distribution ofN-sulfate groups. The characterization of the various disaccharides by Smith periodic acid degradation and glycosidase digestions was facilitated by the preparation and thin-layer chromatographic resolution of the complete series of monosulfated derivatives of anhydromannitol and anhydrotalitol; the sulfate esters were shown to be stable to both the hydrazine and nitrous acid treatments. The high sensitivity of the hydrazine-nitrous acid fragmentation procedure should prove useful in the structural elucidation of cell surface and basement membrane proteoglycans as well as other sulfated glycoconjugates which are present in small amounts.