在拟南芥中表达的海栖热袍菌耐热β-葡萄糖醛酸酶的特性研究。

Characterization of a thermostable β-glucuronidase from Thermotoga maritima expressed in Arabidopsis thaliana.

机构信息

Agro-Biotechnology Research Institute, Shanghai Academy of Agricultural Sciences, 2901 Beidi Rd., Shanghai, 201106, China.

出版信息

Appl Microbiol Biotechnol. 2012 Sep;95(5):1211-9. doi: 10.1007/s00253-011-3802-5. Epub 2011 Dec 24.

DOI:10.1007/s00253-011-3802-5

PMID:22198718

Abstract

TmGUSI, a gene identical to that encoding a thermostable β-glucuronidase in the hyperthermophilic anaerobe Thermotoga maritima, has been synthesized using a PCR-based two-step DNA synthesis and codon optimization for plants, and expressed in both Escherichia coli and Arabidopsis thaliana. TmGUSI expressed in transformed E. coli cells exhibited maximum hydrolytic activity at 65 °C and pH 6.5 and retained more than 80% activity after incubation at 85 °C for 30 min. TmGUSI activity in transgenic A. thaliana plants containing TmGUSI was also stable over the temperature range 65-80 °C. Our data suggest that β-glucuronidase from T. maritima can serve as a useful thermostable marker in higher plants.

摘要

TmGUSI，与嗜热厌氧微生物 Thermotoga maritima 中编码热稳定β-葡萄糖醛酸酶的基因完全相同，已通过基于 PCR 的两步 DNA 合成和针对植物的密码子优化进行了合成，并在大肠杆菌和拟南芥中进行了表达。在转化的大肠杆菌细胞中表达的 TmGUSI 在 65°C 和 pH6.5 时表现出最大的水解活性，在 85°C 孵育 30 分钟后保留超过 80%的活性。含有 TmGUSI 的转基因拟南芥植物中的 TmGUSI 活性在 65-80°C 的温度范围内也很稳定。我们的数据表明，来自 Thermotoga maritima 的β-葡萄糖醛酸酶可以作为高等植物中有用的耐热标记物。