Procopio Noemi, Hopkins Rachel J A, Harvey Virginia L, Buckley Michael
Forensic Science Research Group, Faculty of Health and Life Sciences, Northumbria University, Northumbria University Newcastle, Ellison Building, Newcastle Upon Tyne NE1 8ST, U.K.
Manchester Institute of Biotechnology, The University of Manchester, 131 Princess Street, Manchester M1 7DN, U.K.
J Proteome Res. 2021 Mar 5;20(3):1754-1769. doi: 10.1021/acs.jproteome.0c01014. Epub 2021 Feb 2.
Isotope analyses are some of the most common analytical methods applied to ancient bone, aiding the interpretation of past diets and chronology. For this, the evaluation of "collagen yield" (as defined in radiocarbon dating and stable isotope research) is a routine step that allows for the selection of specimens that are deemed adequate for subsequent analyses, with samples containing less than ∼1% "collagen yield" normally being used for isotopic analysis but discounted for radiocarbon dating. The aims of this study were to use proteomic methods of MALDI-TOF (matrix assisted laser desorption ionization time-of-fligh mass spectrometry) and LC-ESI-MS/MS (liquid chromatography electrospray ionization tandem mass spectrometry) to investigate the endogeneity of the dominant proteinaceous biomolecules within samples that are typically considered to contain poorly preserved protein. Taking 29 archaeological samples, we evaluated the proteome variability between different acid-soluble fractions removed prior to protein gelatinization and considered waste as part of the radiocarbon dating process. We then correlated these proteomes against the commonly used "collagen yield" proxy for preservation. We found that these waste fractions contained a significant amount of both collagenous and noncollagenous proteins (NCPs) but that the abundance of these was not correlated with the acquired "collagen yield". Rather than a depleted protein load as would be expected from a low "collagen yield", the variety of the extracted NCPs was comparable with that commonly obtained from ancient samples and included informative proteins useful for species identification, phylogenetic studies, and potentially even for isotopic analyses, given further method developments. Additionally, we did not observe any correlation between "collagen yield" and peptide mass fingerprint success or between the different fractions taken from the same sample but at different radiocarbon pretreatment stages. Overall, these findings highlight the value in retaining and analyzing sample fractions that are otherwise discarded as waste during the radiocarbon dating process but more importantly, that low "collagen yield" specimens that are often misinterpreted by archaeologists as being devoid of protein can still yield useful molecular sequence-based information.
同位素分析是应用于古代骨骼的一些最常见的分析方法，有助于解读过去的饮食和年代顺序。为此，评估“胶原蛋白产量”（如在放射性碳年代测定和稳定同位素研究中所定义）是一个常规步骤，可用于选择被认为适合后续分析的标本，通常胶原蛋白产量低于约1%的样本用于同位素分析，但不用于放射性碳年代测定。本研究的目的是使用基质辅助激光解吸电离飞行时间质谱（MALDI-TOF）和液相色谱电喷雾电离串联质谱（LC-ESI-MS/MS）等蛋白质组学方法，研究通常被认为蛋白质保存不佳的样本中主要蛋白质生物分子的内源性。我们选取了29个考古样本，评估了在蛋白质凝胶化之前去除的不同酸溶性组分之间的蛋白质组变异性，并将废弃物视为放射性碳年代测定过程的一部分。然后，我们将这些蛋白质组与常用的保存“胶原蛋白产量”指标进行关联。我们发现，这些废弃物组分含有大量的胶原蛋白和非胶原蛋白（NCPs），但它们的丰度与获得的“胶原蛋白产量”无关。与低“胶原蛋白产量”预期的蛋白质负载减少不同，提取的NCPs的种类与通常从古代样本中获得的种类相当，并且包括有助于物种鉴定、系统发育研究的信息性蛋白质，甚至在进一步的方法开发后，可能对同位素分析也有用。此外，我们没有观察到“胶原蛋白产量”与肽质量指纹图谱成功率之间，或来自同一样本但处于不同放射性碳预处理阶段的不同组分之间存在任何相关性。总体而言，这些发现凸显了保留和分析在放射性碳年代测定过程中通常作为废弃物丢弃的样本组分的价值，但更重要的是，那些经常被考古学家误解为不含蛋白质的低“胶原蛋白产量”标本，仍然可以产生基于分子序列的有用信息。
