Bioaffinity Screening with a Rapid and Sample-Efficient Autosampler for Native Electrospray Ionization Mass Spectrometry,Analytical Chemistry

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Bioaffinity Screening with a Rapid and Sample-Efficient Autosampler for Native Electrospray Ionization Mass Spectrometry
Analytical Chemistry ( IF 6.7 ) Pub Date : 2021-09-21 , DOI: 10.1021/acs.analchem.1c03130
Jérôme Kaeslin ₁ , Cyrill Brunner ₁ , Sahar Ghiasikhou ₁ , Gisbert Schneider ₁ , Renato Zenobi ₁

Affiliation

Fast and efficient handling of ligands and biological targets are required in bioaffinity screening based on native electrospray ionization mass spectrometry (ESI-MS). We use a prototype microfluidic autosampler, called the “gap sampler”, to sequentially mix and electrospray individual small molecule ligands together with a target protein and compare the screening results with data from thermal shift assay and surface plasmon resonance. In a first round, all three techniques were used for a screening of 110 ligands against bovine carbonic anhydrase II, which resulted in five mutual hits and some false positives with ESI-MS presumably due to the high ligand concentration or interferences from dimethyl sulfoxide. In a second round, 33 compounds were screened in lower concentrations and in a less complex matrix, resulting in only true positives with ESI-MS. Within a cycle time of 30 s, dissociation constants were determined within an order of magnitude accuracy consuming only 5 pmol of ligand and less than 15 pmol of protein per screened compound. In a third round, dissociation constants of five compounds were accurately determined in a titration experiment. Thus, the gap sampler can rapidly and efficiently be used for high-throughput screening.

中文翻译：

使用快速且样品效率高的自动进样器进行天然电喷雾电离质谱法的生物亲和性筛选

在基于天然电喷雾电离质谱 (ESI-MS) 的生物亲和力筛选中，需要快速有效地处理配体和生物靶标。我们使用原型微流体自动进样器，称为“间隙进样器”，将单个小分子配体与目标蛋白按顺序混合和电喷雾，并将筛选结果与热位移测定和表面等离子体共振的数据进行比较。在第一轮中，所有三种技术都用于针对牛碳酸酐酶 II 筛选 110 个配体，这导致五个相互命中和 ESI-MS 的一些假阳性，这可能是由于高配体浓度或二甲基亚砜的干扰。在第二轮中，以较低浓度和不太复杂的基质筛选了 33 种化合物，使用 ESI-MS 仅产生真正的阳性结果。在 30 秒的循环时间内，每个筛选化合物仅消耗 5 pmol 配体和少于 15 pmol 蛋白质，在一个数量级精度内确定解离常数。在第三轮中，在滴定实验中准确测定了五种化合物的解离常数。因此，间隙采样器可以快速有效地用于高通量筛选。

更新日期：2021-10-06

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