[en] Uroporphyrinogen decarboxylase (UROD) is a key enzyme for the synthesis of chlorophyll, light-sensitive pigment and heme. In order to study the functions of the UROD gene especially in the variations of the leaf color of Cymbidium hybrid, Cymbidium hybrid K21 and its leaf color mutant K21-1 were used as materials, based on the transcriptome sequencing results, ChUROD gene was isolated, its sequence information was analyzed and its expression in different tissues and different varieties were detected by qRT-PCR, furthermore, the related physiological characters were determined. The results showed that ChUROD contains one 1191 bp open reading frame which encoding 396 amino acids. The ChUROD protein has a molecular weight of 43. 78 kD belongs to the URO-D-CIMS-like family. The phylogenetic analysis showed that the ChUROD protein of Cymbidium hybrid was closely related to UROD of Cymbidium sinense. qRT-PCR results indicated that the relative expression of ChUROD gene in leaves of K21 were significantly higher than pseudobulbs and roots, and the relative expression level of ChUROD gene in leaves of K21 was significantly higher than that in K21-1. According to the results of the determination of the UROD enzyme concentration, Urogen III content, Coprogen III content and chlorophyll content, the trend of UROD enzyme concentration was consistent with the relative expression of ChUROD gene. The decarboxylation of Urogen III to produce Coprogen III was hindered and Urogen III was accumulated in yellow leaves of K21-1; the contents of chlorophyll a and chlorophyll b in yellow leaves of K21-1 were significantly lower than that in green leaves. The results suggested the ChUROD gene may play an important role in the variation of the leaf color of K21-1. The results provided a reference basis for the function verification of ChUROD gene and mechanism research of leaf color variation in Cymbidium hybrid. (authors)

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[en] Cotton fibers are unicellular seed trichomes and the largest known plant cells. Fiber morphogenesis in cotton is a complex process involving a large number of genes expressed throughout fiber development process. The expression profiling of five gene families in various cotton tissues was carried out through real time PCR. Expression analysis revealed that transcripts of expansin, tubulin and E6 were elevated from 5 to 20 days post anthesis (DPA) fibers. Three Lipid transfer proteins (LTPs) including LTP1, LTP3, LTP7 exhibited highest expression in 10 - 20 DPA fibers. Transcripts of LTP3 were detected in fibers and non fiber tissues that of LTP7 were almost negligible in non fiber tissues. Sucrose phosphate synthase gene showed highest expression in 10 DPA fibers while sucrose synthse (susy) expressed at higher rate in 5-20 DPA fibers as well as roots. The results reveal that most of fiber related genes showed high expression in 5-20 DPA fibers. Comprehensive expression study may help to determine tissue and stage specificity of genes under study. The study may also help to explore complex process of fiber development and understand the role of these genes in fiber development process. Highly expressed genes in fibers may be transformed in cotton for improvement of fiber quality traits. Genes that were expressed specifically in fibers or other tissues could be used for isolation of upstream regulatory sequences. (author)

[en] In Selenomoans ruminantium, a strictly anaerobic and gram negative bacterium, cadaverine and putrescine are the essential constituents of its peptidoglycan. S. ruminantium does not contain both free and bound types of lipoprotein, but it contains cadaverine as a component of its peptidoglycan. S-adenosylmethionine decarboxylase (SAMDC) is a key enzyme for a synthesis of spermidine and spermine in S. ruminantium. The crude extract of S. ruminantium was preincubated at 100 degrees Celcius and its SAMDC activity was measured by using a "1"4C labeled substrate. We report here on a heat stable SAMDC which is able to withstand a temperature up to 100 degrees Celcius

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[en] α-Amino-β-carboxymuconate-ϵ-semialdehyde decarboxylase (ACMSD) plays an important role in L-tryptophan degradation via the kynurenine pathway. ACMSD forms a homodimer and is functionally inactive as a monomer because its catalytic assembly requires an arginine residue from a neighboring subunit. However, how the oligomeric state and self-association of ACMSD are controlled in solution remains unexplored. In this study, we demonstrate that ACMSD from Pseudomonas fluorescens can self-assemble into homodimer, tetramer, and higher-order structures. Using size-exclusion chromatography coupled with small-angle X-ray scattering (SEC-SAXS) analysis, we investigated the ACMSD tetramer structure, and fitting the SAXS data with X-ray crystal structures of the monomeric component, we could generate a pseudo-atomic structure of the tetramer. This analysis revealed a tetramer model of ACMSD as a head-on dimer of dimers. We observed that the tetramer is catalytically more active than the dimer and is in equilibrium with the monomer and dimer. Substituting a critical residue of the dimer–dimer interface, His-110, altered the tetramer dissociation profile by increasing the higher-order oligomer portion in solution without changing the X-ray crystal structure. ACMSD self-association was affected by pH, ionic strength, and other electrostatic interactions. Alignment of ACMSD sequences revealed that His-110 is highly conserved in a few bacteria that utilize nitrobenzoic acid as a sole source of carbon and energy, suggesting a dedicated functional role of ACMSD's self-assembly into the tetrameric and higher-order structures. Finally, these results indicate that the dynamic oligomerization status potentially regulates ACMSD activity and that SEC-SAXS coupled with X-ray crystallography is a powerful tool for studying protein self-association.

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[en] The Zn(II)-containing carbonic anhydrase (CA) has stimulated much effort into the syntheses of simple model complexes that are designed to mimic the coordination environment of the active site. The Zn(II) in CA is coordinated to three histidine imidazols and a water molecule in a distorted tetrahedral geometry, which is considered to expand to five in a transient manner or by anion inhibitor binding. Recently, X-ray crystal analysis of the SCN^- binding to CA showed the Zn(II) ion in an ill-defined five-coordinate complex with SCN^- and a water bound. Kimura et al. discovered that a tetrahedral Zn(II)-OH₂ triamine complex with 1,5,9-triazacyclododecane(L²) is a good model for the active center of CA. In this complex the pKa of the coordinated water molecule is close to that of CA, and the generated L²-Zn(II)-OH species acts as nucleophile in CA-catalyzing reaction, such as in hydrolysis of activated ester. Additional study on the crystal structure of the SCN^- binding to Zn(II) complex of L² showed a trigonal bipyramidal geometry with an equatorial and an apical Zn(II)-NCS bonds. We also reported the crystal structure of [Zn(L¹)(NCS)][NCS], in which the Zn(II) ion reveals a square pyramidal geometry with an apical thiocyanate nitrogen atom. We report herein the preparation and crystal structure of the Zn(II) complex of L¹ with azide ligands

[en] The zinc coordination in 5-aminolevulinate dehydratase was investigated by extended x-ray absorption fine structure (EXAFS) associated with the zinc K-edge. The enzyme binds 8 mol of zinc/mol of octameric protein, but only four zinc ions seem sufficient for full activity. The authors have undertaken a study on four forms of the enzyme: (a) the eight-zinc native enzyme; (b) the enzyme with only the four zinc sites necessary for full activation occupied; (c) the enzyme with the vacant sites of (b) occupied by four lead ions; (d) the product complex between (b) and porphobilinogen. They have shown that two structurally distinct types of zinc sites are available in the enzyme. The site necessary for activity has an average zinc environment best described by two/three histidines and one/zero oxygen from a group such as tyrosine or a solvent molecule at 2.06 ± 0.02 angstrom, one tyrosine or aspartate at 1.91 ± 0.03 angstrom, and one cysteine sulfur at 2.32 ± 0.03 angstrom with a total coordination of five ligands. The unoccupied site in (b) is dominated by a single contribution of four cysteinyl sulfur atoms at 2.28 ± 0.02 angstrom. Spectra from samples (c) and (d) show only small changes from that of (b), reflecting a slight rearrangement of the ligands around the zinc atom