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Cytochrome P450 monooxygenases play important roles in metabolism. Here, we report the identification and biochemical characterization of P450CHC, a novel self-sufficient cytochrome P450, from cyclohexanecarboxylatedegrading Paraburkholderia terrae KU-64. P450CHC was found to comprise a [2Fe-2S] ferredoxin domain, NAD (P) H-dependent FAD-containing reductase domain, FCD domain, and cytochrome P450 domain ( in that order from the N terminus ) . Reverse transcription--polymerase chain reaction results indicated that the P450 CHC-encoding chcA gene was inducible by cyclohexanecarboxylate. chcA overexpression in Escherichia coli and recombinant protein purification enabled functional characterization of P450CHC as a catalytically self-sufficient cytochrome P450 that hydroxylates cyclohexanecarboxylate. Kinetic analysis indicated that P450 CHC largely preferred NADH (Km = 0.011 m m ) over NADPH (Km = 0.21 mm ). The Kd, Km, and k cat values for cyclohexanecarboxylate were 0.083 m m, 0.084 m m, and 15.9 s-1, respectively. The genetic and biochemical analyses indicated that the physiological role of P450 CHC is initial hydroxylation in the cyclohexanecarboxylate degradation pathway. [ABSTRACT FROM AUTHOR]

The article describes the case of a 45-year-old female patient with a history of talipes equinovarus, fragile skin and hearing impairment, colon diverticulitis and who developed Ehlers-Danlos syndrome (EDS) caused by dermatan-4-Ο-sulfotransferase-1 (D4ST1) deficiency. The diagnosis is confirmed after performing physical examination, microscopic investigations, and genomic DNA sequencing.

The article describes the case of a Japanese boy diagnosed with the symmetrical giant facial plaque variant of juvenile xanthogranuloma (SGFP-JXG), whose condition persisted beyond ten years of age. Topics discussed include JXG's usual manifestation as an asymptomatic nodule that grows slowly, spontaneous resolution that may leave an atrophic scar, and the presence of JXG on the patient's arms. Also mentioned are limited therapeutic options and another case of SGFP-JXG.

Seven xylene-utilizing bacterial strains were isolated from seawater collected off the coast of Japan. Analysis of 16S rRNA gene sequences indicated that six isolates were most closely related to the marine bacterial genera Alteromonas, Marinobacter or Aestuariibacter. The sequence of the remaining strain, KU68FT, showed low similarity to the 16S rRNA gene sequences of known bacteria with validly published names, the most similar species being Maricurvus nonylphenolicus strain KU41ET (92.6% identity). On the basis of physiological, chemotaxonomic and phylogenetic data, strain KU68FT is suggested to represent a novel species of a new genus in the family Cellvibrionaceae of the order Cellvibrionales within the Gammaproteobacteria, for which the name Halioxenophilus aromaticivorans gen. nov., sp. nov. is proposed. The type strain of Halioxenophilus aromaticivorans is KU68FT (=JCM 19134T = KCTC 32387T). PCR and sequence analysis revealed that strain KU68Ft possesses an entire set of genes encoding the enzymes for the upper xylene methyl-monooxygenase pathway, xylCMABN, resembling the gene set of the terrestrial Pseudomonas putida strain mt-2. [ABSTRACT FROM AUTHOR]

The article describes the case of a 50-year-old female patient with a history of subcutaneous mass on her left cheek and who developed angiofibroma of soft tissue (AFST) due to gene rearrangement in the nuclear receptor coactivator 2 gene (NCOA2). The diagnosis was confirmed after performing magnetic resonance imaging (MRI), histopathology, immunihistochemistry and chromogenic in situ hybridization ((CISH). A discussion on the presence of NCOA-2 in tumor cells is presented.

Abstract  Strain KUFI-6N of Exophiala jeanselmei, a cyclohexanol-utilizing yeast-like fungus, was found to grow on 3 isomers of hydroxybenzoate that functioned as the sole carbon sources. Distinct and highly specific hydroxylases converted p- and m-hydroxybenzoate to protocatechuate and o-hydroxybenzoate to catechol. [ABSTRACT FROM AUTHOR]

The enzymatic activity of Pseudomonas fluorescens α-amino-β-carboxymuconic-ϵ-semialdehyde decarboxylase (ACMSD) is critically dependent on a transition metal ion [Li, T., Walker, A. L., Iwaki, H., Hasegawa, Y., and Liu, A. (2005) J. Am. Chem. Soc. 127, 12282-12290]. Sequence analysis in this study further suggests that ACMSD belongs to the amidohydrolase superfamily, whose structurally characterized members comprise a catalytically essential metal cofactor. To identify ACMSD's metal ligands and assess their functions in catalysis, a site-directed mutagenesis analysis was conducted. Alteration of His-9, His-177, and Asp-294 resulted in a dramatic loss of enzyme activity, substantial reduction of the metal-binding ability, and an altered metallocenter electronic structure. Thus, these residues are confirmed to be the endogenous metal ligands. His-11 is implicated in metal binding because of the strictly conserved HxH motif with His-9. Mutations at the 228 site yielded nearly inactive enzyme variants H228A and H228E. The two His-228 mutant proteins, however, exhibited full metal-binding ability and a metal center similar to that of the wild-type enzyme as shown by EPR spectroscopy. Kinetic analysis on the mutants indicates that His-228 is a critical catalytic residue along with the metal cofactor. Since the identified metal ligands and His-228 are present in all known ACMSD sequences, it is likely that ACMSD proteins from other organisms contain the same cofactor and share similar catalytic mechanisms. ACMSD is therefore the first characterized member in the amidohydrolase superfamily that represents a C-C breaking activity. [ABSTRACT FROM AUTHOR]

Pseudomonas fluorescens strain KU-7 is a prototype microorganism that metabolizes 2-nitrobenzoate (2-NBA) via the formation of 3-hydroxyanthranilate (3-HAA), a known antioxidant and reductant. The initial two steps leading to the sequential formation of 2-hydroxy/aminobenzoate and 3-HAA are catalyzed by a NADPH-dependent 2-NBA nitroreductase (NbaA) and 2-hydroxylaminobenzoate mutase (NbaB), respectively. The 216-amino-acid protein NbaA is 78% identical to a plasmid-encoded hypothetical conserved protein of Polaromonas strain JS666; structurally, it belongs to the homodimeric NADH:flavin mononucleotide (FMN) oxidoreductase-like fold family. Structural modeling of complexes with the flavin, coenzyme, and substrate suggested specific residues contributing to the NbaA catalytic activity, assuming a ping-pong reaction mechanism. Mutational analysis supports the roles of Asn40, Asp76, and Glu113, which are predicted to form the binding site for a divalent metal ion implicated in FMN binding, and a role in NADPH binding for the 10-residue insertion in the β5-α2 loop. The 181-amino-acid sequence of NbaB is 35% identical to the 4-hydroxylaminobenzoate lyases (PnbBs) of various 4-nitrobenzoate-assimilating bacteria, e.g., Pseudomonas putida strain TW3. Coexpression of nbaB with nbaA in Escherichia coli produced a small amount of 3-HAA from 2-NBA, supporting the functionality of the nbaB gene. We also showed by gene knockout and chemotaxis assays that nbaY, a chemoreceptor NahY homolog located downstream of the nbaA gene, is responsible for strain KU-7 being attracted to 2-NBA. NbaY is the first chemoreceptor in nitroaromatic metabolism to be identified, and this study completes the gene elucidation of 2-NBA metabolism that is localized within a 24-kb chromosomal locus of strain KU-7. [ABSTRACT FROM AUTHOR]