Structures and Mechanisms of Two Enzymes that Produce Hydrocarbon Fuels from Abundant Metabolites

有了这个奖项，“化学部”的“生命过程的化学”项目正在支持宾夕法尼亚州立大学的工作，研究两种催化脂肪酸（丰富和常见的细胞代谢产物）转化为碳氢化合物（烷烃和烯烃）的关键步骤的酶。然后，这些可以用作内燃机的直接生物燃料。支持的研究正在影响可再生能源的发展，因为微生物的使用是生物燃料生产的一种有前途的策略。这个过程特别环保，因为它最终是碳中和的，并由太阳能驱动。该项目正在研究上述酶如何发挥作用，以期将它们导入能够生产生物燃料的基因工程微生物。 该奖项支持的工作也为学生和博士后学者提供了在生物物理方法方面接受广泛培训的机会。它还支持由宾夕法尼亚州立大学金属生物化学家领导并由其他机构的同事支持的两年一度的研讨会，该研讨会培训学生和博士后学者在该研究领域使用的独特方法。可以（i）从阳光和二氧化碳中产生脂肪酸（直接通过使用光合蓝细菌，或间接通过向工程微生物饲喂从快速生长和高产植物如甘蔗获得的糖）和（ii）将脂肪酸转化为“直接”生物燃料的遗传可操纵微生物被积极地用作可再生燃料的生物发生的载体。最近发现的两种用于将脂肪酸转化为直链烷烃或烯烃的酶途径涉及在其活性位点具有含铁辅因子的酶。第一个例子涉及蓝细菌中的两步烷烃生物合成途径，其通过脂肪醛将脂肪酸转化为烷（a/e）。该途径的第二步是由铁蛋白样二铁羧酸氧化酶和加氧酶的一员--腺苷脱甲酰化加氧酶（ADO）催化的。在上一个资助周期中，我们研究了ADO反应作为一种新的，神秘的氧化还原加氧反应。进一步的机制研究揭示了潜在的低效率和脆弱性，可能会限制这一途径对生物燃料生产的有用性。我们建议更深入地研究ADO的反应，期望进一步的机理洞察力将指导生物燃料生产微生物的合理设计。第二个例子涉及最近发现的铁酶UndA，它利用O2将脂肪酸脱羧为二氧化碳和末端烯烃。我们的初步工作表明，UndA也是一种二铁酶，与已发表的假设相反，它是一种单核铁酶。我们建议确定正确的辅因子，并确定详细的分子机制。这些研究将对正在进行的旨在利用这种代谢途径生产生物燃料的研究工作做出重要贡献。

With this award the "Chemistry of Life Processes" program of the "Division of Chemistry" is supporting work at the Pennsylvania State University to study two enzymes that catalyze key steps in the conversion of fatty acids (abundant and common cellular metabolites) to hydrocarbons (alkanes and alkenes). These can then be used as drop-in biofuels for combustion engines. The research supported is impacting development of renewable energy, because the use of microorganisms is a promising strategy for biofuel production. This process is particularly environmentally friendly, because it is ultimately carbon neutral and driven by solar energy. The project is studying how the enzymes described above function with an eye to importing them into genetically engineered microorganisms which can produce biofuels. Work supported by this award is also offering students and postdoctoral scholars opportunities to be broadly trained in biophysical methods. It also supports a biennial workshop, led by Penn State metallobiochemists and supported by colleagues from other institutions, that trains students and postdoctoral scholars in unique methods used in this research field. Genetically manipulable microorganisms that could (i) produce fatty acids from sunlight and carbon dioxide (either directly by using photosynthetic cyanobacteria, or indirectly by feeding the engineered microorganisms sugars obtained from fast-growing and high-yielding plants such as sugar cane) and (ii) convert the fatty acids into "drop-in" biofuels are actively pursued as vehicles for biogenesis of renewable fuels. Two recently discovered enzyme pathways for the conversion of fatty acids into linear alkane or alkene hydrocarbons involve enzymes that harbor iron-containing cofactors at their active site. The first example involves the two-step alkane biosynthesis pathway in cyanobacteria that converts fatty acids via fatty aldehydes to alk(a/e)nes. The second step in this pathway is catalyzed by the enzyme aldehyde-deformylating oxygenase (ADO), a member of the ferritin-like diiron-carboxylate oxidases and oxygenases. In the previous funding cycle we studied the ADO reaction as a novel, cryptically redox oxygenation reaction. Further mechanistic studies revealed potential inefficiencies and vulnerabilities that could limit the usefulness of this pathway for biofuel production. We propose to study the reaction of ADO in greater depth, with the expectation that further mechanistic insight will guide the rational design of biofuel-producing microorganisms. The second example involves a recently discovered iron-enzyme, UndA, that utilizes O2 to decarboxylate fatty acids to carbon dioxide and terminal alkenes. Our preliminary work suggests that UndA is also a diiron enzyme, contrary to the published assumption that it is a mononuclear iron enzyme. We propose to identify the correct cofactor and determine the molecular mechanism in detail. These studies will be an important contribution to ongoing research efforts that aim at utilizing this metabolic pathway for biofuel production.