title: A novel flavin-containing monooxygenase from Methylophaga sp strain SK1 and its indigo synthesis in Escherichia coli.
abstract: We cloned a gene from Methylophaga sp. strain SK1. This gene was responsible for producing, the blue pigment, indigo. The complete open reading frame was 1371 bp long, which encodes a protein of 456 amino acids. The molecular mass of the encoded protein was 105 kDa, consisting of homodimer of 54 kDa with an isoelectric point of 5.14. The deduced amino acid sequence from the gene showed approximately 30% identities with flavin-containing monooxygenases (FMOs) of human (FMO1-FMO5), Arabidopsis, and yeast. It contained three characteristic sequence motifs of FMOs: FAD binding domain, FMO-identifying sequence motif, and NADPH binding domain. In addition, the biochemical properties such as substrate specificities and absorption spectra were similar to the eukaryotic FMO families. Thus, we assigned the enzyme to be a bacterial FMO. The recombinant Escherichia coli expressing the bacterial FMO produced up to 160 mg of indigo per liter in the tryptophan medium after 12h cultivation. This suggests that the recombinant strain has a potential to be applied in microbial indigo production.
authors.affiliation: Research Center for Proteineous Materials, Chosun University, Gwangju 501-759, Republic of Korea.
authors.forename: Hack Sun
authors.initials: HS
authors.lastname: Choi
authors.forename: Jin Kwon
authors.initials: JK
authors.lastname: Kim
authors.forename: Eun Hee
authors.initials: EH
authors.lastname: Cho
authors.forename: Yong Chul
authors.initials: YC
authors.lastname: Kim
authors.forename: Jae Il
authors.initials: JI
authors.lastname: Kim
authors.forename: Si Wouk
authors.initials: SW
authors.lastname: Kim
mesh_terms: D020816:Amino Acid Motifs; D000595:Amino Acid Sequence; D002851:Chromatography, High Pressure Liquid; D003001:Cloning, Molecular; D019281:Dimerization; D004926:Escherichia coli / Q000378:metabolism*; D005182:Flavin-Adenine Dinucleotide / Q000378:metabolism; D005415:Flavins / Q000737:chemistry*; D005656:Fungal Proteins / Q000737:chemistry; D020563:Gammaproteobacteria / Q000201:enzymology*; D015723:Gene Library; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007525:Isoelectric Focusing; D007700:Kinetics; D006899:Mixed Function Oxygenases / Q000737:chemistry*; D008969:Molecular Sequence Data; D016297:Mutagenesis, Site-Directed; D009249:NADP / Q000378:metabolism; D016366:Open Reading Frames; D010860:Pigments, Biological / Q000096:biosynthesis*; D010940:Plant Proteins / Q000737:chemistry; D010957:Plasmids / Q000378:metabolism; D017434:Protein Structure, Tertiary; D017386:Sequence Homology, Amino Acid; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D013053:Spectrophotometry; D013379:Substrate Specificity; D013997:Time Factors
chemical_list: D005415:Flavins; D005656:Fungal Proteins; D010860:Pigments, Biological; D010940:Plant Proteins; D005182:Flavin-Adenine Dinucleotide; D009249:NADP; D006899:Mixed Function Oxygenases