Development of a Cold-Adapted Pseudoalteromonas Expression System for the Pseudoalteromonas Proteins Intractable for the Escherichia coli System

PloS One
Zi-Chao YuYu-Zhong Zhang

Abstract

Although the Escherichia coli expression system is the most commonly used expression system, some proteins are still difficult to be expressed by this system, such as proteins with high thermolability and enzymes that cannot mature by autoprocessing. Therefore, it is necessary to develop alternative expression systems. In this study, a cold-adapted Pseudoalteromonas expression system was developed. A shuttle vector was constructed, and a conjugational transfer system between E. coli and psychrophilic strain Pseudoalteromonas sp. SM20429 was established. Based on the shuttle vector, three reporter vectors were constructed to compare the strength of the cloned promoters at low temperature. The promoter of xylanase gene from Pseudoalteromonas sp. BSi20429 was chosen due to its high activity at 10-15°C. An expression vector pEV containing the chosen promoter, multiple cloning sites and a His tag was constructed for protein expression and purification. With pEV as expression vector and SM20429 as the host, a cold-adapted protease, pseudoalterin, which cannot be maturely expressed in E. coli, was successfully expressed as an active extracellular enzyme when induced by 2% oat spelt xylan at 15°C for 48 h. Recombinant pseudoalterin pur...Continue Reading

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Citations

Dec 29, 2015·Bioengineered·Gro Elin Kjæreng BjergaAdele Williamson
Sep 27, 2016·Frontiers in Microbiology·Margarita SantiagoLoreto P Parra
Sep 7, 2019·Natural Product Reports·Yannick BuijsSheng-Da Zhang
Nov 13, 2019·Microbial Cell Factories·Jenny Johansson SöderbergPeik Haugen
Jan 17, 2020·Nature Communications·Bai-Lu TangYu-Zhong Zhang

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Datasets Mentioned

BETA
ET12567
BADV00000000

Methods Mentioned

BETA
PCR

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