重組蛋白質在大腸桿菌之異體表現情況與條件的探討

Abstract

蛋白質是生物體的重要組成之一，可以調控或催化許多的生化反應。但是，在傳統的蛋白質製程上，大多經由生體進行萃取取得蛋白質，受到流程繁瑣與產量少的困擾，所以造成取得不易且成本過高。因此，透過使用大腸桿菌、酵母菌或病毒等等的微生物異體表現系統，並利用蛋白質的物化性質，把目標蛋白純化出來，來取得不易或成本過高的蛋白質，是當前生產蛋白質藥物的重要策略之一。 本研究的目的在觀察與探討蛋白質在異體表達宿主大腸桿菌中的表現性與可溶情形，並瞭解用不同的純化方式，對蛋白質表現與純度的影響。實驗中有改變大腸桿菌培養條件，以及誘導蛋白質表現的誘導劑濃度，來瞭解培養條件對其表現的影響，另外也把伴隨蛋白一起放進大腸桿菌中，來瞭解對蛋白質可溶性程度的影響，後續將可溶的粗抽蛋白液進行親和性樹脂的純化，以瞭解管柱沖提條件對重組蛋白的純化情況的影響。

Proteins are important components of living organisms and can regulate or catalyze many biochemical reactions. However, the traditional procedure by extraction to obtain the proteins often suffers from complication and low production yield. In recent years, heterologous expression system has been developed for better efficiency in production. Some common microorganisms are used for the heterologous expression, such as E. coli and yeast. The protein products from such expression system can then be purified according to their physical or chemical properties. This study observes and discusses the expression level of a model recombinant protein by changing conditions of expression and incubation of E. coli. In a few cases, a chaperone can be tested to help the solubility of the protein. An attempt was also made to examine the purification condition of the protein by affinity chromatography.