[Cloning, expression and characterization of alcohol dehydrogenase and aldehyde dehydrogenase from Bacillus pseudofirmus OF4].

[Cloning, expression and characterization of alcohol dehydrogenase and aldehyde dehydrogenase from Bacillus pseudofirmus OF4].

Wei sheng wu xue bao = Acta microbiologica Sinica (2013-07-19)

Jiansong Ju, Ning Ma, Ranran Zhao, Jingwei Liu, Shujing Xu, Baohua Zhao

摘要

To clone and characterize the alcohol dehydrogenase (ADH) and aldehyde dehydrogenase (ALDH) from Bacillus pseudofirmus OF4. Genes of adh and aldh were cloned by PCR; expression vectors pET-Ahd and pET-Aldh were constructed and expressed in Escherichia coli BL21 (DE3). After Ni-NTA column chromatography purification, the protein was characterized. The optimal temperature and pH of ALDH was 35 degrees C and 8.0, the specific activities of ALDH was 979.6 U/mg protein, the thermostability at 25 degrees C and 35 degrees C was better than at 45 degrees C. Although the expression level of ADH was too low to purify, but it was found that ADH had high catalytic activities by experiments of co-expression and ethanol tolerance. Adh and aldh from B. pseudofirmus OF4 were cloned successfully. Co-expression of double genes could greatly increase the host strain on ethanol tolerance.

材料

货号

品牌

产品描述

E7023

Sigma-Aldrich

乙醇，Pure 200纯度

459844

Sigma-Aldrich

纯乙醇, 200 proof, ACS reagent, ≥99.5%

459828

Sigma-Aldrich

纯乙醇, 200 proof

493546

Sigma-Aldrich

纯乙醇, 200 proof, meets USP testing specifications

E7148

Sigma-Aldrich

乙醇，Pure 190纯度, for molecular biology

02870

Sigma-Aldrich

酒精, ACS reagent, prima fine spirit, without additive, F15 o¹

270741

Sigma-Aldrich

酒精试剂, denatured, suitable for HPLC

362808

Sigma-Aldrich

酒精试剂, reagent grade

493511

Sigma-Aldrich

纯乙醇, 190 proof, ACS spectrophotometric grade, 95.0%

02857

Sigma-Aldrich

酒精, purum, fine spirit, denaturated with 4.8% methanol, F25 METHYL¹, ~96% (based on denaturant-free substance)

E-029

Supelco

乙醇-50标准液 CRM, 50 mg/dL in H₂O, ampule of 10 × 1.2 mL, certified reference material, Cerilliant^®

459836

Sigma-Aldrich

纯乙醇, 200 proof, anhydrous, ≥99.5%

E-040

Supelco

乙醇-10标准液 CRM, 10 mg/dL in H₂O, pack of 10 × 1.2 mL ampules, certified reference material, Cerilliant^®

48075

Supelco

Ethanol 溶液, certified reference material, 2000 μg/mL in methanol

32221

Sigma-Aldrich

酒精, puriss. p.a., absolute, ≥99.8% (GC)

E-030

Supelco

乙醇-80 标准液 CRM, 80 mg/dL in H₂O, ampule of 10 × 1.2 mL, certified reference material, Cerilliant^®

E-053

Supelco

乙醇-500标准液 CRM, 500 mg/dL in H₂O, ampule of 10 × 1.2 mL, certified reference material, Cerilliant^®

E-031

Supelco

乙醇-100（10支/盒)标准液 CRM, 100 mg/dL in H₂O, ampule of 10 × 1.2 mL, certified reference material, Cerilliant^®

E-033

Supelco

乙醇-300标准液 CRM, 300 mg/dL in H₂O, ampule of 10 × 1.2 mL, certified reference material, Cerilliant^®

E2385

Supelco

10% (v/v) 乙醇标准品, 10 % (v/v) in H₂O, analytical standard

02882

Sigma-Aldrich

酒精, purum, absolute ethanol, denaturated with 2% 2-butanone, A15 MEK¹, ≥99.8% (based on denaturant-free substance)

E-041

Supelco

乙醇-150, 150 mg/dL in H₂O, ampule of 10 × 1.2 mL, certified reference material, Cerilliant^®

09-0851

Sigma-Aldrich

酒精, ≥99.5%, suitable for HPLC

E-032

Supelco

乙醇-200 (10支/盒)标准液 CRM, 200 mg/dL in H₂O, ampule of 10 × 1.2 mL, certified reference material, Cerilliant^®

09-0790

Sigma-Aldrich

酒精, JIS special grade, 94.8-95.8%

09-0852

Sigma-Aldrich

酒精, ≥99.5%

E-056

Supelco

乙醇-20（10支/盒)标准液 CRM, 20 mg/dL in H₂O, ampule of 10 × 1.2 mL, certified reference material, Cerilliant^®

E-035

Supelco

Ethanol 溶液, 25 mg/dL in H₂O, ampule of 10 × 1.2 mL, certified reference material, Cerilliant^®

E-036

Supelco

乙醇-400（10支/盒)标准液 CRM, 400 mg/dL in H₂O, ampule of 10 × 1.2 mL, certified reference material, Cerilliant^®

E-034

Supelco

乙醇校准套装, ampule of 10 × 1.2 mL, certified reference material, Cerilliant^®