Purification and Characterization of the Coniferyl Aldehyde Dehydrogenase from Pseudomonas sp. Strain HR199 and Molecular Characterization of the Gene

Sandra Achterholt; Horst Priefert; Alexander Steinbüchel

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Purification and Characterization of the Coniferyl Aldehyde Dehydrogenase from Pseudomonas sp. Strain HR199 and Molecular Characterization of the Gene

机译：假单胞菌属的针叶树醛醛脱氢酶的纯化和鉴定。 HR199菌株和该基因的分子表征

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The coniferyl aldehyde dehydrogenase (CALDH) ofPseudomonas sp. strain HR199 (DSM7063), which catalyzes the NAD+-dependent oxidation of coniferyl aldehyde to ferulic acid and which is induced during growth with eugenol as the carbon source, was purified and characterized. The native protein exhibited an apparent molecular mass of 86,000 ± 5,000 Da, and the subunit mass was 49.5 ± 2.5 kDa, indicating an α₂ structure of the native enzyme. The optimal oxidation of coniferyl aldehyde to ferulic acid was obtained at a pH of 8.8 and a temperature of 26°C. The K_m values for coniferyl aldehyde and NAD+ were about 7 to 12 μM and 334 μM, respectively. The enzyme also accepted other aromatic aldehydes as substrates, whereas aliphatic aldehydes were not accepted. The NH₂-terminal amino acid sequence of CALDH was determined in order to clone the encoding gene (calB). The corresponding nucleotide sequence was localized on a 9.4-kbp EcoRI fragment (E94), which was subcloned from a Pseudomonas sp. strain HR199 genomic library in the cosmid pVK100. The partial sequencing of this fragment revealed an open reading frame of 1,446 bp encoding a protein with a relative molecular weight of 51,822. The deduced amino acid sequence, which is reported for the first time for a structural gene of a CALDH, exhibited up to 38.5% amino acid identity (60% similarity) to NAD+-dependent aldehyde dehydrogenases from different sources.

机译：假单胞菌（Pseudomonas） sp。的针叶树醛脱氢酶（CALDH）。纯化并表征了菌株HR199（DSM7063），该菌株催化松柏基醛的NAD + 依赖性氧化成阿魏酸，并在以丁香酚为碳源的生长过程中被诱导。天然蛋白的表观分子量为86,000±5,000 Da，亚单位质量为49.5±2.5 kDa，表明天然酶的α_{2 结构。在8.8的pH值和26°C的温度下，针叶树醛醛向阿魏酸的最佳氧化作用。松柏醛和NAD + 的 K _{m 值分别约为7至12μM和334μM。该酶还接受其他芳香族醛作为底物，而脂肪族醛则不被接受。测定CALDH的NH _{2 末端氨基酸序列，以克隆编码基因（ calB ）。相应的核苷酸序列位于一个9.4kbp的 Eco RI片段（E94）上，该片段亚克隆自假单胞菌 sp。粘粒pVK100中的HR199基因组文库。该片段的部分测序揭示了一个1,446 bp的开放阅读框，编码一个相对分子量为51,822的蛋白质。首次报道的CALDH结构基因推导的氨基酸序列与来自不同来源的NAD + 依赖的醛脱氢酶具有高达38.5％的氨基酸同一性（60％相似性）。资料来源。}}} 展开▼

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《Journal of bacteriology》 |1998年第17期|共5页

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Sandra Achterholt; Horst Priefert; Alexander Steinbüchel;
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中图分类微生物学;

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专利

1. A coniferyl aldehyde dehydrogenase gene from Pseudomonas sp strain HR199 enhances the conversion of coniferyl aldehyde by Saccharomyces cerevisiae [J] . Adeboye Peter Temitope, Olsson Lisbeth, Bettiga Maurizio Bioresource Technology: Biomass, Bioenergy, Biowastes, Conversion Technologies, Biotransformations, Production Technologies . 2016,第Null期

机译：假单胞菌属菌株HR199的针叶树醛脱氢酶基因增强了酿酒酵母对针叶树醛的转化

2. Identification and molecular characterization of the eugenol hydroxylase genes (ehyA/ehyB) of Pseudomonas sp. strain HR199 [J] . Horst Priefert, Jorg Overhage, Alexander Steinbuchel Archives of microbiology . 1999,第6期

机译：假单胞菌sp。的丁香酚羟化酶基因（ehyA / ehyB）的鉴定和分子表征。 HR199株

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6. Purification and Characterization of the Coniferyl Aldehyde Dehydrogenase from Pseudomonas sp. Strain HR199 and Molecular Characterization of the Gene [O] . Sandra Achterholt, Horst Priefert, Alexander Steinbüchel 1998

机译：假单胞菌属的针叶树醛醛脱氢酶的纯化和鉴定。 HR199菌株和该基因的分子表征

7. A coniferyl aldehyde dehydrogenase gene from Pseudomonas sp. strain HR199 enhances the conversion of coniferyl aldehyde by Saccharomyces cerevisiae [O] . Adeboye, Peter Temitope, Olsson, Lisbeth, Bettiga, Maurizio 2016

机译：来自假单胞菌属（Pseudomonas sp。）的针叶树醛脱氢酶基因。菌株HR199增强酿酒酵母对松柏醛的转化。

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机译：菌株组成包括：嗜麦芽单胞菌2L菌株，5L嗜麦芽单胞菌sp菌株，嗜麦芽单胞菌sp菌株6L，嗜麦芽单胞菌sp菌株3N，无色杆菌属菌株4P，节杆菌属菌株1N，短杆菌属菌株2N，短杆菌属菌株，短杆菌属菌株5N，短杆菌属菌株6N，短杆菌属sp菌株3G，4G菌株假单胞菌sp。保藏下保藏号为KKP 2041 p，包含生物修复疫苗菌株的成分，利用疫苗对土壤进行生物修复以去除污染物和土壤净化方法

5. Strain composition comprising: Stenotrophomonas sp strain 2L, 5L Stenotrophomonas sp strain, Stenotrophomonas sp strain 6L, Stenotrophomonas sp strain 3N, Achromobacter sp strain 4P, Arthrobacter sp strain 1N, 2N Brevundimonas sp strain, Brevundimonas sp strain 5N, 6N Brevuedimonas sp strain, Pseudomonas sp strain 3G, 4G strain Pseudomonas sp. deposited under - deposit number KKP 2041 p, bioremediation vaccine strains containing the composition, the use of vaccine bioremediation of soil to remove contaminants and soil purification method [P] . 外国专利： PL399388A1 . 2013-12-09

机译：菌株组成包括：嗜麦芽单胞菌2L菌株，5L嗜麦芽单胞菌sp菌株，嗜麦芽单胞菌sp菌株6L，嗜麦芽单胞菌sp菌株3N，无色杆菌属菌株4P，节杆菌属菌株1N，短杆菌属菌株2N，短杆菌属菌株，短杆菌属菌株5N，短杆菌属菌株6N，短杆菌属sp菌株3G，4G菌株假单胞菌sp。保藏下保藏号为KKP 2041 p，包含生物修复疫苗菌株的成分，利用疫苗对土壤进行生物修复以去除污染物和土壤净化方法

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Purification and Characterization of the Coniferyl Aldehyde Dehydrogenase from Pseudomonas sp. Strain HR199 and Molecular Characterization of the Gene

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