Journal
FERMENTATION-BASEL
Volume 8, Issue 10, Pages -Publisher
MDPI
DOI: 10.3390/fermentation8100498
Keywords
alpha-glucosidase; isomaltooligosaccharides; isopeptide bonds; thermal stability
Funding
- National Key Research and Development Program of China [2021YFC2100900]
- National Natural Science Foundation of China [32171471, 32071470]
- Key Research and Development Project of Shandong Province, China [2019JZZY020605]
- Priority Academic Program Development of Jiangsu Higher Education Institutions
- Top-notch Academic Programs Project of Jiangsu Higher Education Institutions
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The thermal stability of alpha-glucosidase was significantly improved by constructing cyclized proteins, allowing for continuous production of isomaltooligosaccharides at higher temperatures.
alpha-glucosidase is an essential enzyme for the production of isomaltooligosaccharides (IMOs). Allowing alpha-glucosidase to operate at higher temperatures (above 60 degrees C) has many advantages, including reducing the viscosity of the reaction solution, enhancing the catalytic reaction rate, and achieving continuous production of IMOs. In the present study, the thermal stability of alpha-glucosidase was significantly improved by constructing cyclized proteins. We screened a thermotolerant alpha-glucosidase (AGL) with high transglycosylation activity from Thermoanaerobacter ethanolicus JW200 and heterologously expressed it in Bacillus subtilis 168. After forming the cyclized alpha-glucosidase by different isopeptide bonds (SpyTag/SpyCatcher, SnoopTag/SnoopCatcher, SdyTag/SdyCatcher, RIAD/RIDD), we determined the enzymatic properties of cyclized AGL. The optimal temperature of all cyclized AGL was increased by 5 degrees C, and their thermal stability was generally improved, with SpyTag-AGL-SpyCatcher having a 1.74-fold increase compared to the wild-type. The results of molecular dynamics simulations showed that the RMSF values of cyclized AGL decreased, indicating that the rigidity of the cyclized protein increased. This study provides an efficient method for improving the thermal stability of alpha-glucosidase.
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