Deciphering stable isotope fractionation (d13C; d37Cl) during reductive dehalogenation of chlorinated ethenes: Effects of bacterial growth physiology and expression of key enzymes

解析氯化乙烯还原脱卤过程中的稳定同位素分馏（d13C；d37Cl）：细菌生长生理学和关键酶表达的影响

• 批准号: 250925090
• 负责人: Professor Dr. Sebastian Felix Behrens, since 9/2014
• 金额: --
• 依托单位: University of Minnesota
• 依托单位国家: 德国
• 项目类别: Research Grants
• 财政年份: 2013
• 资助国家: 德国
• 起止时间: 2012-12-31 至 2017-12-31
• 项目状态: 已结题
• 来源: https://gepris.dfg.de/gepris/projekt/250925090?language=en
• 关键词: Deciphering; stable; isotope; fractionation; d13C

Compound specific isotope analysis (CSIA) is regarded as a powerful approach to assess in situ biotransformation of chlorinated organic compounds as based on kinetic isotope fractionation due to biodegradation. Although reported carbon (d13C) isotope fractionation factors for microbial reductive dehalogenation of chlorinated ethenes vary considerably even for the same bacterial strain and thus hamper the interpretation and applicability of isotope data in complex environmental settings, systematic experimental investigations on factors and mechanisms influencing observed isotope fractionation are scarce. The major objective of our proposed research is to identify and evaluate what and how physiologic and enzymatic factors control isotope fractionation during microbial reductive dechlorination of chlorinated ethenes. Generally, the extent of stable isotope fractionation may be determined largely by the reaction mechanism at the respective enzyme, but can be attenuated by uptake and transport processes of the substrate. More specifically, controlling factors may include concentrations and type of electron donor/acceptor, availability of growth supplements (vitamins/trace elements), growth phase, cell density or expression and abundance of reductive dehalogenase genes. The majority of the known and genetically characterized organohalide reducing bacteria (OHRBs) carry multiple homologous genes encoding putative reductive dehalogenases. In order to directly link observed dehalogenation activity and isotope fractionation to gene expression levels we propose Desulfitobacterium hafniense strain Y51 as model organism for our study because it possesses only two key enzymes that are both well characterized (PCE and chlorophenol reductive dehalogenases). This model organism will thus facilitate a systematic evaluation and discrimination of various potential effects of selected growth conditions on isotope fraction in conjunction with molecular analyses of specific gene expression and protein content. Our recently developed expertise in compound specific chlorine (d37Cl) isotope analysis enables us to determine also chlorine isotope fractionation factors for microbial dehalogenation. Thus, through improved mechanistic understanding of the controlling factors of microbial isotope fractionation the proposed research will enhance the applicability and practical value of 2-dimensional carbon/chlorine isotope approaches to assess in situ biodegradation of organochlorine compounds in the field and will provide an experimental basis for linking compound specific isotope data with reactive transport models.

化合物特异性同位素分析（CSIA）是一种基于动力学同位素分馏的评价氯化有机化合物在生物降解过程中原位生物转化的有效方法。尽管报道的氯化乙烯微生物还原脱卤的碳（d13C）同位素分馏因子即使在同一菌株中也存在很大差异，从而妨碍了同位素数据在复杂环境下的解释和适用性，但对影响观察到的同位素分馏的因素和机制的系统实验研究很少。我们提出的研究的主要目的是确定和评估生理和酶的因素是什么，以及如何控制在微生物还原脱氯氯化乙烯过程中的同位素分馏。一般来说，稳定同位素分离的程度很大程度上取决于相应酶的反应机制，但可以通过底物的吸收和运输过程减弱。更具体地说，控制因素可能包括电子供体/受体的浓度和类型、生长补充剂（维生素/微量元素）的可用性、生长阶段、细胞密度或还原脱卤酶基因的表达和丰度。大多数已知的和遗传特征的有机卤化物还原细菌（OHRBs）携带多个同源基因编码推定的还原脱卤酶。为了将观察到的脱卤活性和同位素分离与基因表达水平直接联系起来，我们提出了hafniense Desulfitobacterium Y51菌株作为我们研究的模式生物，因为它只有两种关键酶（PCE和氯酚还原脱卤酶）。因此，该模式生物将结合特定基因表达和蛋白质含量的分子分析，促进对选定生长条件对同位素分数的各种潜在影响的系统评估和区分。我们最近在化合物特定氯（d37Cl）同位素分析方面的专业知识使我们能够确定微生物脱卤的氯同位素分馏因素。因此，通过提高对微生物同位素分异控制因素的机理理解，本研究将增强二维碳/氯同位素方法在现场评估有机氯化合物原位生物降解的适用性和实用价值，并将为将化合物特定同位素数据与反应输运模型联系起来提供实验基础。