Sriraman Kalpana, Jayaraman Guhan
Department of Biotechnology, Indian Institute of Technology-Madras, Chennai 600036, India.
Appl Microbiol Biotechnol. 2006 Oct;72(6):1202-9. doi: 10.1007/s00253-006-0410-x. Epub 2006 Apr 1.
Lactococcus lactis is a potential host for production of recombinant proteins, especially of therapeutic importance. However, in glucose-grown cultures, lowering of pH due to accumulation of lactic acid and the concomitant induction of acid tolerance response (ATR) may affect the recombinant protein produced. In this work, we have analyzed the effect of culture pH and the associated ATR on production of recombinant streptokinase. Streptokinase gene was cloned and expressed as a secretory protein in L. lactis under the control of P170 promoter. It was found to undergo degradation to form inactive products leading to low productivity. The extent of degradation and productivity of streptokinase was greatly influenced by the development of ATR, which was dependent on the pH of the culture and initial phosphate concentration of the medium. It was found that high pH and high initial phosphate concentration leads to suppression of ATR and this results in at least 2.5-fold increase in streptokinase productivity and significant decrease in degradation of streptokinase.
乳酸乳球菌是生产重组蛋白，尤其是具有治疗重要性的重组蛋白的潜在宿主。然而，在以葡萄糖为生长底物的培养物中，由于乳酸积累导致的pH降低以及随之而来的耐酸反应（ATR）诱导可能会影响所产生的重组蛋白。在这项工作中，我们分析了培养pH值和相关的ATR对重组链激酶生产的影响。链激酶基因被克隆，并在P170启动子的控制下作为分泌蛋白在乳酸乳球菌中表达。结果发现它会发生降解形成无活性产物，导致生产率低下。链激酶的降解程度和生产率受到ATR发展的极大影响，而ATR又取决于培养物的pH值和培养基的初始磷酸盐浓度。结果发现，高pH值和高初始磷酸盐浓度会抑制ATR，这导致链激酶生产率至少提高2.5倍，并且链激酶的降解显著减少。
