Mas Sandrine, Villas-Bôas Silas Granato, Hansen Michael Edberg, Akesson Mats, Nielsen Jens
Center for Microbial Biotechnology, BioCentrum-DTU, Technical University of Denmark, Building 223, DK-2800 Kgs. Lyngby, Denmark.
Biotechnol Bioeng. 2007 Apr 1;96(5):1014-22. doi: 10.1002/bit.21194.
Recent technical advances in mass spectrometry (MS) have propelled this technology to the forefront of methods employed in metabolome analysis. Here, we compare two distinct analytical approaches based on MS for their potential in revealing specific metabolic footprints of yeast single-deletion mutants. Filtered fermentation broth samples were analyzed by GC-MS and direct infusion ESI-MS. The potential of both methods in producing specific and, therefore, discriminant metabolite profiles was evaluated using samples from several yeast deletion mutants grown in batch-culture conditions with glucose as the carbon source. The mutants evaluated were cat8, gln3, ino2, opi1, and nil1, all with deletion of genes involved in nutrient sensing and regulation. From the analysis, we found that both methods can be used to classify mutants, but the classification depends on which metabolites are measured. Thus, the GC-MS method is good for classification of mutants with altered nitrogen regulation as it primarily measures amino acids, whereas this method cannot classify mutants involved in regulation of phospholipids metabolism as well as the direct infusion MS (DI-MS) method. From the analysis, we find that it is possible to discriminate the mutants in both the exponential and stationary growth phase, but the data from the exponential growth phase provide more physiological relevant information. Based on the data, we identified metabolites that are primarily involved in discrimination of the different mutants, and hereby providing a link between high-throughput metabolome analysis, strain classification, and physiology.
质谱（MS）技术的最新进展已将该技术推向代谢组分析所采用方法的前沿。在此，我们比较基于质谱的两种不同分析方法在揭示酵母单基因缺失突变体特定代谢特征方面的潜力。通过气相色谱 - 质谱联用（GC - MS）和直接进样电喷雾电离质谱（ESI - MS）对过滤后的发酵液样品进行分析。使用在以葡萄糖为碳源的分批培养条件下生长的几种酵母缺失突变体的样品，评估这两种方法生成特定且具有判别力的代谢物谱的潜力。所评估的突变体有cat8、gln3、ino2、opi1和nil1，所有这些突变体的相关基因缺失均涉及营养感知和调控。通过分析，我们发现这两种方法均可用于对突变体进行分类，但分类取决于所测量的代谢物。因此，GC - MS方法适用于对氮调节改变的突变体进行分类，因为它主要测量氨基酸，而该方法对涉及磷脂代谢调控的突变体的分类效果不如直接进样质谱（DI - MS）方法。通过分析，我们发现有可能在指数生长期和稳定期对突变体进行区分，但指数生长期的数据提供了更多与生理相关的信息。基于这些数据，我们鉴定出主要参与区分不同突变体的代谢物，从而在高通量代谢组分析、菌株分类和生理学之间建立了联系。
