A CALCULATION OF ALL POSSIBLE OLIGOSACCHARIDE ISOMERS BOTH BRANCHED AND LINEAR YIELDS 1.05X10(12) STRUCTURES FOR A REDUCING HEXASACCHARIDE - THE ISOMER-BARRIER TO DEVELOPMENT OF SINGLE-METHOD SACCHARIDE SEQUENCING OR SYNTHESIS SYSTEMS
A CALCULATION OF ALL POSSIBLE OLIGOSACCHARIDE ISOMERS BOTH BRANCHED AND LINEAR YIELDS 1.05X10(12) STRUCTURES FOR A REDUCING HEXASACCHARIDE - THE ISOMER-BARRIER TO DEVELOPMENT OF SINGLE-METHOD SACCHARIDE SEQUENCING OR SYNTHESIS SYSTEMS
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DOI：
10.1093/glycob/4.6.759
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发表时间：
1994-12-01
期刊：
影响因子：
4.3
通讯作者：
LAINE, RA
中科院分区：
文献类型：
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作者：
LAINE, RA
计算了六糖的所有可能的线性和支链异构体的数量，发现> 1.05 x 10（12）。这个大的数字定义了异构体屏障，这是开发用于碳水化合物的绝对表征的单一分析方法的持久技术屏障，无论样品量如何，由于这种异构体屏障，没有单一的方法可以用于确定100 nmol量的完整寡糖结构，程序Edman肽或桑格DNA测序方法。在开发寡糖的简易合成方案中的困难也可以通过这个大的数量来解释，目前没有化学或物理分析方法具有区分具有相同质量的10（12）个结构所必需的分辨率。因此，仅通过NMR或质谱法对中等重量低聚糖进行“表征”必然包含非常大的误差范围。如一些商业广告所吹捧的，仅通过顺序酶降解然后凝胶渗透色谱法或电泳法进行的结果具有更大的不确定性。因此，许多使用这些单一方法来“表征”复杂碳水化合物的文献都存在问题，期刊应该注意发表结构表征，除非作者揭示了所有可能的替代结构。今天，只有定量糖分析，甲基化连接分析，通过酶或化学的部分降解和质谱的组合才能将可能性的数量减少到一个。本研究产生了一些单独的公式和一组主方程，这些公式和方程对于确定二糖到八糖的所有可能的还原末端异构体是必要的，高于这些公式和方程的分支异构体产生天文数字，因为六糖通常是最大的生物活性、蛋白质识别的寡糖序列，并且也是多糖中最大的重复单元，所以本计算限于dp 6。尽管有这种简化，但计算的用于还原仅由D己糖组成的六糖的可能结构的数量> 10（12）。生物活性低聚糖的可用微量化学需要10至100 nmol，以实现采用部分降解的湿化学/酶学/质谱法的最低必要组合。尽管进行了大量的研究活动，但碳水化合物（与蛋白质和DNA相比）分析的相对较高的限制量20年来一直保持不变。该计算强调了在碳水化合物分析中开发与Edman蛋白和桑格DNA测序方法在灵敏度上相当的微量化学的长期技术障碍的原因，它还揭示了与用于肽合成的那些开发的方法相当的用于寡糖的简易合成方法的障碍。
The number of all possible linear and branched isomers of a hexasaccharide was calculated and found to be > 1.05 x 10(12). This large number defines the Isomer Barrier, a persistent technological barrier to the development of a single analytical method for the absolute characterization of carbohydrates, regardless of sample quantity, Because of this isomer barrier, no single method can be employed to determine complete oligosaccharide structure in 100 nmol amounts with the same assurance that can be achieved for 100 pmol amounts with single-procedure Edman peptide or Sanger DNA sequencing methods. Difficulties in the development of facile synthetic schemes for oligosaccharides are also explained by this large number, No current method of chemical or physical analysis has the resolution necessary to distinguish among 10(12) structures having the same mass. Therefore the 'characterization' of a middle-weight oligosaccharide solely by NMR or mass spectrometry necessarily contains a very large margin of error, Greater uncertainty accompanies results performed solely by sequential enzyme degradation followed by gel-permeation chromatography or electrophoresis, as touted by some commercial advertisements. Much of the literature which uses these single methods to 'characterize' complex carbohydrates is, therefore, in question, and journals should beware of publishing structural characterizations unless the authors reveal all alternate possible structures which could result from their analysis. Today, only a combination of quantitative sugar analysis, methylation linkage analysis, partial degradation by enzymes or chemistry, and mass spectrometry can reduce the number of possibilities to one, The present study yields a number of individual formulae and a master set of equations necessary for the determination of all possible reducing-end isomers for di- to octasaccharides, above which branching isomers generate astronomical numbers, larger than Avogadro's number, Because hexasaccharides are generally among the largest biologically active, protein-recognized oligosaccharide sequences, and also among the largest repeating units in polysaccharides, the present calculation was limited to dp6. Despite this simplification, the number of possible structures calculated for reducing hexasaccharides comprised of D hexoses alone is > 10(12). Available microchemistry for biologically active oligosaccharides requires between 10 and 100 nmol for a minimum necessary combination of wet chemistry/enzymology/mass spectrometry employing partial degradation. The relatively high limiting quantity for analysis of carbohydrates (compared with proteins and DNA) has remained static for 20 years, despite intense research activity. This calculation underscores the reason for the long-standing technology barrier for the development of a microchemistry in carbohydrate analysis comparable in sensitivity with Edman protein and Sanger DNA sequencing methods, It also reveals the barrier to facile synthetic methods for oligosaccharides comparable to those developed for peptide synthesis.