Evaluation of the function of a luciferase-like monooxygenase homologue in 4,4´-dithiodibutyric acid catabolism in Rhodococcus erythropolis MI2  

作　　者：M.Venkateswar Reddy Alexander Steinbüchel 

机构地区：[1]Institut für Molekulare Mikrobiologie und Biotechnologie,Westfälische Wilhelms-Universität Münster,48149 Münster,Germany [2]Center for Biotechnology and Interdisciplinary Studies,Rensselaer Polytechnic Institute,1108th Street,Troy,NY 12180,USA [3]Environmental Sciences Department,King Abdulaziz University,Jeddah,Saudi Arabia

出　　处：《Systems Microbiology and Biomanufacturing》2022年第3期523-532,共10页系统微生物学与生物制造（英文）

基　　金：supported by Alexander von Humboldt(AvH)foundation,Germany(Ref No:IND 1162665 HFST-P)。

摘　　要：The bacterium Rhodococcus erythropolis MI2 uses 4,4´-dithiodibutyric acid(DTDB)as carbon source to synthesize polythioesters(PTE).The first step for the production of PTE using DTDB is catalyzed by an NADH:flavin oxidoreductase(nox)as it was previously shown in our laboratory,and the second step is catabolized by a putative luciferase-like monooxygenase(Llm).In the current study,experiments were carried out to identify the function of Llm.Hence,the llm gene,which encodes for the Llm protein,was amplified from the genomic DNA of MI2 using polymerase chain reaction and expressed in Escherichia coli BL21 cells.Protein purification was done using His Spin Trap affinity columns.Enzyme assay was carried out using the purified protein and p-coumaric acid as substrate giving a specific activity of 1.6 U/mg protein for the purified Llm.The responsible gene(llm)was deleted in the genome of MI2,and a single deletion mutant was subsequently generated.Finally,growth of the wild-type strain(MI2)and the mutant strain(MI2Δllm)were compared using DTDB or succinate as carbon sources.Whereas the wild type was successfully grown with DTDB or succinate,the llm-negative mutant exhibited low grow with DTDB although it grows very well with succinate.

关 键 词：Deletion mutant MONOOXYGENASE Polythioesters Protein purification Rhodococcus erythropolis MI2 4 4´-Dithiodibutyric acid(DTDB) 

分 类 号：TL5[核科学技术—核技术及应用]