茂源链霉菌(Streptomyces mobaraensis)谷氨酰胺转氨酶(transglutaminase,TGase)活化蛋白酶(activating metalloprotease,TAMEP)能够专一性切割TGase的酶原区,其高效制备对于重组TGase的生产有重要意义。将S. mobaraensis 来源的TAMEP成功表达于大肠杆菌。通过对信号肽类型、信号肽N端密码子及诱导表达条件的优化,使胞外TAMEP酶活力达到186.3 U/mL。酶学性质分析显示,重组TAMEP的最适反应温度和最适反应pH分别为55 ℃和7.0;TAMEP在30~50 ℃孵育60 min酶活力保持在50%以上,且在pH 6.2~8.9较稳定;0.133 μmol/L重组 TAMEP能够在30 min内使6.91 μmol/L TGase基本活化。研究结果为TAMEP规模化制备提供了生产菌株和基础数据。
Streptomyces mobaraensis transglutaminase (TGase)-activating metalloprotease (TAMEP) can specifically cleave the zymogen region of TGase to form the active form. In this study,TAMEP from S. mobaraensis was successfully expressed in Escherichia coli. Through the optimizations of signal peptide type,N-terminal codon of the signal peptide,and induction conditions,the extracellular activity of TAMEP reached 186.3 U/mL. As indicated by enzymatic analysis,that the optimal reaction temperature and pH of the recombinant TAMEP were 55 ℃ and pH 7.0,respectively. 50% activity of TAMEP was retained after incubating at 30-50 ℃ for 60 min,and the enzyme was also stable at pH 6.2-8.9. 0.133 μmol/L recombinant TAMEP can basically activated 6.91 μmol/L TGase in 30 minutes. The research results provided strains and basic data for the large-scale preparation of TAMEP.
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