Perera Madushika, Wijesundera Sulochana, Wijayarathna C Dilrukshi, Seneviratne Gamini, Jayasena Sharmila
Department of Biochemistry and Molecular Biology, Faculty of Medicine, University of Colombo, Colombo, Sri Lanka.
Department of Chemistry, Faculty of Science, University of Colombo, Colombo, Sri Lanka.
Front Microbiol. 2022 Aug 30;13:898456. doi: 10.3389/fmicb.2022.898456. eCollection 2022.
Efficient degradation of alkanes in crude oil by the isolated MM1 alluded to the presence of highly active alkane-degrading enzymes in this fungus. A long-chain alkane-degrading, LadA-like enzyme family in was identified, and possible substrate-binding modes were analyzed using a computational approach. By analyzing publicly available protein databases, we identified six uncharacterized proteins in NRRL 3357, of which five were identified as class LadAα and one as class LadAβ, which are eukaryotic homologs of bacterial long-chain alkane monooxygenase (LadA). Computational models of LadAα homologs () showed overall structural similarity to the bacterial LadA and the unique sequence and structural elements that bind the cofactor Flavin mononucleotide (FMN). A receptor-cofactor-substrate docking protocol was established and validated to demonstrate the substrate binding in the LadAα homologs. The modeled , , and captured long-chain -alkanes inside the active pocket, above the bound FMN. Isoalloxazine ring of reduced FMN formed a π-alkyl interaction with the terminal carbon atom of captured alkanes, C-C, in 3-5 and C-C in 1. Our results confirmed the ability of identified LadAα monooxygenases to bind long-chain alkanes inside the active pocket. Hence . LadAα monooxygenases potentially initiate the degradation of long-chain alkanes by oxidizing bound long-chain alkanes into their corresponding alcohol.
分离得到的MM1能高效降解原油中的烷烃，这表明该真菌中存在高活性的烷烃降解酶。在该真菌中鉴定出了一个类似LadA的长链烷烃降解酶家族，并采用计算方法分析了可能的底物结合模式。通过分析公开的蛋白质数据库，我们在NRRL 3357中鉴定出6个未表征的蛋白质，其中5个被鉴定为LadAα类，1个被鉴定为LadAβ类，它们是细菌长链烷烃单加氧酶（LadA）的真核同源物。LadAα同源物的计算模型显示，其整体结构与细菌LadA相似，且具有结合辅因子黄素单核苷酸（FMN）的独特序列和结构元件。建立并验证了一种受体-辅因子-底物对接方案，以证明LadAα同源物中的底物结合情况。模拟的[具体编号1]、[具体编号2]和[具体编号3]在结合的FMN上方的活性口袋内捕获了长链烷烃。还原型FMN的异咯嗪环与捕获的烷烃的末端碳原子形成了π-烷基相互作用，在[具体编号3]中为C-C，在[具体编号1]中为C-C。我们的结果证实了鉴定出的LadAα单加氧酶在活性口袋内结合长链烷烃的能力。因此，LadAα单加氧酶可能通过将结合的长链烷烃氧化为相应的醇来启动长链烷烃的降解。
