胶原蛋白是结缔组织中一种重要蛋白质,具有抗皱、抗老化和补钙壮骨的功效,在食品工业中应用广泛。由于胶原蛋白的需求量大,因此利用重组微生物生产胶原蛋白成为近年来的研究热点。该文采用重组有类人胶原蛋白基因的毕赤酵母,通过优化培养基成分,使其在无机盐培养基中表达胶原蛋白。首先,类人胶原蛋白在每24 h补加体积分数1.5%甲醇诱导120 h后的产量可达18.87 mg/L,进而将无机盐培养基中盐浓度降至1/3,有效避免了类人胶原蛋白的水解,也减少了重组毕赤酵母菌株分泌的杂蛋白量。进一步添加1 g/L L-脯氨酸等辅助营养物使类人胶原蛋白的产量提高至40.26 mg/L,为初始类人胶原蛋白表达量的2倍。结果表明,添加氨基酸等氮源能显著提高重组毕赤酵母的类人胶原蛋白分泌量,为重组类人胶原蛋白的发酵生产提供了指导。
Collagen is an important structural protein in connective tissue, plays an important role in anti-wrinkle, anti-aging and calcium reinforcement, and widely applies in food industry. This research optimized the cultivation condition of recombinant Komagataella phaffii to express human-like collagen in an inorganic salt medium. Firstly, the yield of human-like collagen was 18.87 mg/L at 1.5% methanol induction per 24 h after 120 h induced fermentation. Further, degradation of human-like collagen was avoided by reducing the salts concentration to 1/3 of initial value, also leading to less impure protein in fermentation broth. The yield of human-like collagen reached 40.26 mg/L at the condition 1 g/L L-proline addition, which was 2-fold of initial yield of human-like collagen. This research indicated that the nitrogen source, such as amino acid, increased the yield of human-like collagen by recombinant K. phaffii significantly, which provides an important direction for collagen production through fermentation.
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