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Jeerus Sucharitakul Chanakan Tongsook Danaya Pakotiprapha Willem J. H. van Berkel Pimchai Chaiyen 《The Journal of biological chemistry》2013,288(49):35210-35221
3-Hydroxybenzoate 6-hydroxylase (3HB6H) from Rhodococcus jostii RHA1 is an NADH-specific flavoprotein monooxygenase that catalyzes the para-hydroxylation of 3-hydroxybenzoate (3HB) to form 2,5-dihydroxybenzoate (2,5-DHB). Based on results from stopped-flow spectrophotometry, the reduced enzyme-3HB complex reacts with oxygen to form a C4a-peroxy flavin with a rate constant of 1.13 ± 0.01 × 106
m−1 s−1 (pH 8.0, 4 °C). This intermediate is subsequently protonated to form a C4a-hydroperoxyflavin with a rate constant of 96 ± 3 s−1. This step shows a solvent kinetic isotope effect of 1.7. Based on rapid-quench measurements, the hydroxylation occurs with a rate constant of 36 ± 2 s−1. 3HB6H does not exhibit substrate inhibition on the flavin oxidation step, a common characteristic found in most ortho-hydroxylation enzymes. The apparent kcat at saturating concentrations of 3HB, NADH, and oxygen is 6.49 ± 0.02 s−1. Pre-steady state and steady-state kinetic data were used to construct the catalytic cycle of the reaction. The data indicate that the steps of product release (11.7 s−1) and hydroxylation (36 ± 2 s−1) partially control the overall turnover. 相似文献
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Genotypic variation in milling depression of iron and zinc concentration in rice grain 总被引:1,自引:0,他引:1
Saenchai Chorpet Prom-u-thai Chanakan Jamjod Sansanee Dell Bernard Rerkasem Benjavan 《Plant and Soil》2012,360(1-2):271-286
Plant and Soil - The loss of iron and zinc during milling to produce white rice can have serious consequences for human health. Therefore, the objective was to evaluate Fe and Zn partitioning... 相似文献
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David Secco Nadia Bouain Aida Rouached Chanakan Prom-u-thai Moez Hanin Ajay K. Pandey 《Critical reviews in biotechnology》2017,37(7):898-910
Phosphorus (P) is an essential macronutrient for all living organisms. In plants, P is taken up from the rhizosphere by the roots mainly as inorganic phosphate (Pi), which is required in large and sufficient quantities to maximize crop yields. In today’s agricultural society, crop yield is mostly ensured by the excessive use of Pi fertilizers, a costly practice neither eco-friendly or sustainable. Therefore, generating plants with improved P use efficiency (PUE) is of major interest. Among the various strategies employed to date, attempts to engineer genetically modified crops with improved capacity to utilize phytate (PA), the largest soil P form and unfortunately not taken up by plants, remains a key challenge. To meet these challenges, we need a better understanding of the mechanisms regulating Pi sensing, signaling, transport and storage in plants. In this review, we summarize the current knowledge on these aspects, which are mainly gained from investigations conducted in Arabidopsis thaliana, and we extended it to those available on an economically important crop, wheat. Strategies to enhance the PA use, through the use of bacterial or fungal phytases and other attempts of reducing seed PA levels, are also discussed. We critically review these data in terms of their potential for use as a technology for genetic manipulation of PUE in wheat, which would be both economically and environmentally beneficial. 相似文献
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Tonapha Pusadee Chanakan Prom-u-thai Narit Yimyam Sansanee Jamjod Benjavan Rerkasem 《Economic botany》2017,71(2):175-187
Phenotypic and Genetic Diversity of Local Perilla ( Perilla frutescens (L.) Britt.) from Northern Thailand. Perilla frutescens (L.) Britt., an important oil and culinary crop in Asia, is a valuable genetic resource. Despite its nutritional value and historic and cultural importance, research on Perilla has been scarce, particularly as far as its genetic diversity is concerned. The aims of the present study were to assess variability within and between 29 seed samples of P. frutescens collected from farmers in northern Thailand, and evaluation conducted of their genetic, morphological, and agronomic characteristics, and the seed composition, including polyunsaturated fatty acids omega-3, omega-6, and omega-9, and the vitamin E γ-tocopherols. Perilla frutescens (L.) Britt. of northern Thailand is genetically variable, and structured according to origin of collection which was the consequence of local adaptation. The discovery of high levels of polyunsaturated fatty acids, namely α-linolenic acid and γ-tocopherols, in some Perilla samples indicates the potential for utilizing Perilla for its high omega-3 content including as a vitamin E supplement for humans, a prospect that should be taken into account when planning conservation strategies or when Perilla variability is used in breeding programs. 相似文献
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Tongsook C Sucharitakul J Thotsaporn K Chaiyen P 《The Journal of biological chemistry》2011,286(52):44491-44502
p-Hydroxyphenylacetate (HPA) 3-hydroxylase is a two-component flavoprotein monooxygenase that catalyzes the hydroxylation of p-hydroxyphenylacetate to form 3,4-dihydroxyphenylacetate. Based on structures of the oxygenase component (C2), both His-120 and Ser-146 are located ∼2.8 Å from the hydroxyl group of HPA. The variants H120N, H120Q, H120Y, H120D, and H120E can form C4a-hydroperoxy-FMN (a reactive intermediate necessary for hydroxylation) but cannot hydroxylate HPA. The impairment of H120N is not due to substrate binding because the variant can still bind HPA. In contrast, the H120K variant catalyzes hydroxylation with efficiency comparable with that of the wild-type enzyme; the hydroxylation rate constant for H120K is 5.7 ± 0.6 s−1, and the product conversion ratio is 75%, compared with values of 16 s−1 and 90% for the wild-type enzyme. H120R can also catalyze hydroxylation, suggesting that a positive charge on residue 120 can substitute for the hydroxylation function of His-120. Because the hydroxylation reaction of wild-type C2 is pH-independent between pH 6 and 10, the protonation status of key components required for hydroxylation likely remains unchanged in this pH range. His-120 may be positively charged for selective binding to the phenolate form of HPA, i.e. to form the Hisδ+·HPAδ− complex, which in turn promotes oxygen atom transfer via an electrophilic aromatic substitution mechanism. Analysis of Ser-146 variants revealed that this residue is necessary for but not directly engaged in hydroxylation. Product formation in S146A is pH-independent and constant at ∼70% over a pH range of 6–10, whereas product formation for S146C decreased from ∼65% at pH 6.0 to 27% at pH 10.0. These data indicate that the ionization of Cys-146 in the S146C variant has an adverse effect on hydroxylation, possibly by perturbing formation of the Hisδ+·HPAδ− complex needed for hydroxylation. 相似文献
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Ruangchan N Tongsook C Sucharitakul J Chaiyen P 《The Journal of biological chemistry》2011,286(1):223-233
p-Hydroxyphenylacetate (HPA) 3-hydroxylase (HPAH) catalyzes the hydroxylation of HPA at the ortho-position to yield 3,4-dihydroxyphenylacetate. The enzyme is a flavin-dependent two-component monooxygenase that consists of a reductase component and an oxygenase component (C(2)). C(2) catalyzes the hydroxylation of HPA using oxygen and reduced FMN as co-substrates. To date, the effects of pH on the oxygenation of the two-component monooxygenases have never been reported. Here, we report the reaction kinetics of C(2)·FMNH(-) with oxygen at various pH values investigated by stopped-flow and rapid quenched-flow techniques. In the absence of HPA, the rate constant for the formation of C4a-hydroperoxy-FMN (~1.1 × 10(6) m(-1)s(-1)) was unaffected at pH 6.2-9.9, which indicated that the pK(a) of the enzyme-bound reduced FMN was less than 6.2. The rate constant for the following H(2)O(2) elimination step increased with higher pH, which is consistent with a pK(a) of >9.4. In the presence of HPA, the rate constants for the formation of C4a-hydroperoxy-FMN (~4.8 × 10(4) m(-1)s(-1)) and the ensuing hydroxylation step (15-17 s(-1)) were not significantly affected by the pH. In contrast, the following steps of C4a-hydroxy-FMN dehydration to form oxidized FMN occurred through two pathways that were dependent on the pH of the reaction. One pathway, dominant at low pH, allowed the detection of a C4a-hydroxy-FMN intermediate, whereas the pathway dominant at high pH produced oxidized FMN without an apparent accumulation of the intermediate. However, both pathways efficiently catalyzed hydroxylation without generating significant amounts of wasteful H(2)O(2) at pH 6.2-9.9. The decreased accumulation of the intermediate at higher pH was due to the greater rates of C4a-hydroxy-FMN decay caused by the abolishment of substrate inhibition in the dehydration step at high pH. 相似文献
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