title: Functional analysis of a novel repressor LuxR in Comamonas testosteroni.
abstract: Comamonas testosteroni (C. testosteroni) ATCC11996 is a gram negative bacterium which can use steroid as a carbon and energy source. 3,17β-hydroxysteroid dehydrogenase (3,17β-HSD) is a key enzyme for the degradation of steroid hormones in C. testosteroni. The LuxR regulation family is a group of regulatory proteins which play important role in gram negative bacterium. The luxr gene is located on 58 kb upstream of 3,17β-HSD gene with the opposite transcription orientation in the chromosomal DNA of C. testosteroni. An open reading frame of this putative luxr gene consists of 1125 bp and is translated into a protein containing 374 amino acids. The luxr gene was cloned into plasmid pK18 and plasmid pK-LuxR1 was obtained. E. coli HB101 was co-transformed by pK-LuxR1 and pUC912-10, pUC1128-5 or pUC3.2-4 (which contain βhsd gene and different length promoter, repeat sequences). The result of ELISA showed that LuxR protein is a negative regulator for 3,17β-HSD expression. The luxr gene in C. testosteroni was knock-out by homologous integration. 3,17β-HSD expression was increased in the mutant (C.T.-L-KO1) comparing to that in wild-type C. testosteroni (C.T.) after 0.5 mM testosterone induction. The mutant C.T.-L-KO1 and wild-type C. testosteroni were cultured at 27 °C and 37 °C. The result of growth curve proved that LuxR has also effect on the bacterial growth.
authors.affiliation: Changchun University of Science and Technology, 7989 Weixing Road, 130022 Changchun, China.
authors.forename: Ye
authors.initials: Y
authors.lastname: Ji
authors.affiliation: Department of Family Medicine, The First Affiliated Hospital, School of Medicine, Zhejiang University, 310000 Hangzhou, China; Institute of Toxicology and Pharmacology for Natural Scientists, Medical School Schleswig-Holstein, 24103 Kiel, Germany.
authors.forename: Tianyuan
authors.initials: T
authors.lastname: Pan
authors.affiliation: Changchun University of Science and Technology, 7989 Weixing Road, 130022 Changchun, China.
authors.forename: Yang
authors.initials: Y
authors.lastname: Zhang
authors.affiliation: Institute of Toxicology and Pharmacology for Natural Scientists, Medical School Schleswig-Holstein, 24103 Kiel, Germany.
authors.forename: Guangming
authors.initials: G
authors.lastname: Xiong
authors.affiliation: Changchun University of Science and Technology, 7989 Weixing Road, 130022 Changchun, China. Electronic address: yuyuanhua8888@126.com.
authors.forename: Yuanhua
authors.initials: Y
authors.lastname: Yu
mesh_terms: D015067:17-Hydroxysteroid Dehydrogenases / Q000235:genetics / Q000378:metabolism; D000595:Amino Acid Sequence; D003001:Cloning, Molecular; D020604:Comamonas testosteroni / Q000187:drug effects / Q000201:enzymology* / Q000254:growth & development; D004926:Escherichia coli / Q000378:metabolism; D055786:Gene Knockout Techniques; D010957:Plasmids / Q000235:genetics / Q000378:metabolism; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000737:chemistry / Q000302:isolation & purification; D012097:Repressor Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D013696:Temperature; D013739:Testosterone / Q000494:pharmacology; D015534:Trans-Activators / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D015854:Up-Regulation / Q000187:drug effects
chemical_list: D011994:Recombinant Proteins; D012097:Repressor Proteins; D015534:Trans-Activators; C064778:LuxR autoinducer binding proteins; D013739:Testosterone; D015067:17-Hydroxysteroid Dehydrogenases; C044256:3 (or 17)-beta-hydroxysteroid dehydrogenase
keywords: 3,17β-Hydroxysteroid dehydrogenase; Comamonas testosteroni; Gene regulation; Knock-out mutant; LuxR repressor; Testosterone induction