Hashemi Payam, Luckau Luise, Mischnick Petra, Schmidt Sarah, Stosch Rainer, Wünsch Bettina
Institute for Food Chemistry, Technische Universität Braunschweig, Schleinitzstr. 20, 38106, Braunschweig, Germany.
Metrology in Chemistry, Physikalisch-Technische Bundesanstalt, Bundesallee 100, 38116, Braunschweig, Germany.
Anal Bioanal Chem. 2017 Oct;409(25):5901-5909. doi: 10.1007/s00216-017-0554-9. Epub 2017 Aug 14.
Nucleic acids, proteins, and polysaccharides are the most important classes of biopolymers. The inherent properties of biomacromolecules are contrary to those of well-defined small molecules consequently raising a number of specific challenges which become particularly apparent if biomacromolecules are treated as objects in quantitative analysis. At the same time, their specific functional ability of molecular recognition and self-organization (e.g., enzymes, antibodies, DNA) enables us to make biomacromolecules serving as molecular tools in biochemistry and molecular biology, or as precisely controllable dimensional platforms for nanometrological applications. Given the complexity of biomacromolecules, quantitative analysis is not limited to the measurement of their concentration but also involves the determination of numerous descriptors related to structure, interaction, activity, and function. Among the biomacromolecules, glycans set examples that quantitative characterization is not necessarily directed to the measurement of amount-of-substance concentration but instead involves the determination of relative proportions (molar ratios) of structural features for comparison with theoretical models. This article addresses current activities to combine optical techniques such as Raman spectroscopy with isotope dilution approaches to realize reference measurement procedures for the quantification of protein biomarkers as an alternative to mass spectrometry-based techniques. Furthermore, it is explored how established ID-MS protocols are being modified to make them applicable for quantifying virus proteins to measure the HIV viral load in blood samples. As an example from the class of carbohydrates, the challenges in accurate determination of substitution patterns are outlined and discussed. Finally, it is presented that biomacromolecules can also serve as tools in quantitative measurements of dimensions with an example of DNA origami to generate defined dimensional standards to be used for calibration in super-resolution fluorescence microscopy. Graphical abstract Quantitative analysis of biomacromolecules is accompanied with special challenges different from low molecular weight compounds. In addition, they are not only objects but also tools applicable for quantitative measurements.
核酸、蛋白质和多糖是最重要的生物聚合物类别。生物大分子的固有特性与结构明确的小分子相反，因此带来了一些特殊挑战，若将生物大分子作为定量分析的对象，这些挑战就会变得尤为明显。同时，它们具有分子识别和自组装的特定功能能力（如酶、抗体、DNA），这使我们能够将生物大分子用作生物化学和分子生物学中的分子工具，或用作纳米计量应用中可精确控制尺寸的平台。鉴于生物大分子的复杂性，定量分析不仅限于测量其浓度，还涉及确定与结构、相互作用、活性和功能相关的众多描述符。在生物大分子中，聚糖给出了实例，即定量表征不一定针对物质的量浓度的测量，而是涉及确定结构特征的相对比例（摩尔比），以便与理论模型进行比较。本文介绍了当前将拉曼光谱等光学技术与同位素稀释方法相结合的活动，以实现蛋白质生物标志物定量的参考测量程序，作为基于质谱技术的替代方法。此外，还探讨了如何修改既定的同位素稀释质谱协议，使其适用于定量病毒蛋白以测量血样中的HIV病毒载量。作为碳水化合物类别的一个例子，概述并讨论了准确确定取代模式方面的挑战。最后，通过DNA折纸术的例子展示了生物大分子也可作为尺寸定量测量的工具，以生成用于超分辨率荧光显微镜校准的确定尺寸标准。图形摘要生物大分子的定量分析伴随着与低分子量化合物不同的特殊挑战。此外，它们不仅是分析对象，也是适用于定量测量的工具。
