Journal
BIOTECHNOLOGY FOR BIOFUELS
Volume 9, Issue -, Pages -Publisher
BIOMED CENTRAL LTD
DOI: 10.1186/s13068-016-0490-5
Keywords
SpyTag/SpyCatcher; Cyclization; Non-chromatographic purification; Lichenase; Thermal stability
Funding
- National Natural Science Foundation of China [21376103]
- Promotion Program for Young and Middle-aged Teacher in Science and Technology Research of Huaqiao University [ZQN-YX105]
Ask authors/readers for more resources
Background: SpyTag is a peptide that can form an irreversible covalent linkage to its 12 kDa partner SpyCatcher via a spontaneous isopeptide bond. Herein, we fused SpyTag at the N-terminal of lichenase and SpyCatcher at C-terminal so that the termini of lichenase were locked together by the covalent interaction between the partners. In addition, an elastin-like polypeptides tag was subsequently attached to the C-terminus of SpyCatcher, thereby facilitating the non-chromatographic purification of cyclized lichenase. Results: The study showed that the optimum temperature of the cyclized lichenase was about 5 degrees C higher in comparison to its linear counterpart. Moreover, nearly 80 % of the cyclized lichenase activities were retained after 100 degrees C exposure, whereas the linear form lost almost all of its activities. Therefore, the cyclized variant displayed a significantly higher thermal stability as temperature elevated and was resistant to hyperthermal denaturation. Besides, the Km value of the cyclized lichenase (7.58 +/- 0.92 mg/mL) was approximately 1.7-fold lower than that of the linear one (12.96 +/- 1.93 mg/mL), indicating a higher affinity with substrates. Conclusions: This new SpyTag/SpyCatcher cyclization strategy is deemed as a generalized reference for enhancing enzyme stability and can be effectively customized to the cyclization of various enzymes, hence a tremendous potential for successful application in the biocatalytic conversion of biomass to produce fuels and chemicals.
Authors
I am an author on this paper
Click your name to claim this paper and add it to your profile.
Reviews
Recommended
No Data Available