Manuell Andrea L, Beligni Maria Verónica, Elder John H, Siefker David T, Tran Miller, Weber Annika, McDonald Thomas L, Mayfield Stephen P
The Department of Cell Biology and The Skaggs Institute for Chemical Biology, La Jolla, CA 92037, USA.
Plant Biotechnol J. 2007 May;5(3):402-12. doi: 10.1111/j.1467-7652.2007.00249.x. Epub 2007 Mar 15.
We have engineered the chloroplast of eukaryotic algae to produce a number of recombinant proteins, including human monoclonal antibodies, but, to date, have achieved expression to only 0.5% of total protein. Here, we show that, by engineering the mammalian coding region of bovine mammary-associated serum amyloid (M-SAA) as a direct replacement for the chloroplast psbA coding region, we can achieve expression of recombinant protein above 5% of total protein. Chloroplast-expressed M-SAA accumulates predominantly as a soluble protein, contains the correct amino terminal sequence and has little or no post-translational modification. M-SAA is found in mammalian colostrum and stimulates the production of mucin in the gut, acting in the prophylaxis of bacterial and viral infections. Chloroplast-expressed and purified M-SAA is able to stimulate mucin production in human gut epithelial cell lines. As Chlamydomonas reinhardtii is an edible alga, production of therapeutic proteins in this organism offers the potential for oral delivery of gut-active proteins, such as M-SAA.
我们已对真核藻类的叶绿体进行改造，以生产多种重组蛋白，包括人单克隆抗体，但迄今为止，重组蛋白的表达量仅占总蛋白的0.5%。在此，我们表明，通过对牛乳腺相关血清淀粉样蛋白（M-SAA）的哺乳动物编码区进行改造，直接替换叶绿体psbA编码区，我们能够使重组蛋白的表达量超过总蛋白的5%。叶绿体表达的M-SAA主要以可溶性蛋白形式积累，具有正确的氨基末端序列，且几乎没有或没有翻译后修饰。M-SAA存在于哺乳动物初乳中，可刺激肠道中粘蛋白的产生，起到预防细菌和病毒感染的作用。叶绿体表达并纯化的M-SAA能够刺激人肠道上皮细胞系中粘蛋白的产生。由于莱茵衣藻是一种可食用藻类，在这种生物体中生产治疗性蛋白为口服递送肠道活性蛋白（如M-SAA）提供了可能性。
