蛋白質為生物體中功能之執行者，參與了所有生命活動的過程，其如何執行缜密之功能與調控，為許多研究者致力探討的問題。生化與結構分析為探討蛋白質作用機制的重要手段，但其中仰賴大量且高純度之蛋白質樣本進行分析。然而高等生物之蛋白表現時，往往需要正確的轉譯後修飾或特定的伴蛋白幫助摺疊以行使其功能，這使得生化與結構之研究往往受限於蛋白之取得。近年來隨著基因工程技術的成熟，低成本且具有一定修飾能力之植物表現系統非常具有研究潛力，其中以農桿菌為基因傳遞媒介於植物進行蛋白表現之系統，能在 2 至 6 天內進行蛋白的瞬時表現，可望成為克服過去生化與結構研究瓶頸之平台。因此，本篇研究以 GFP 螢光蛋白建立農桿菌瞬時表現與純化流程，並成功應用此流程取得具生物活性之 AtKAI2並完成生化之分析。在過往不易取得之植物膜蛋白上，藉由目前建立的實驗流程也已可以順利地完成 VrVPPase 之純化並進行負染單分子分析。總合上述結果，農桿菌介導之瞬時表現系統具備高度潛力能應用於進行各種植物蛋白分子之生化與結構研究，進而探討這些植物蛋白在植株中的調控方式與作用機轉。

Proteins serve as the functional molecular machines, play crucial roles in various biological processes in cells. Therefore, studying how proteins sophisticatedly perform their functions is highly attractive to scientists. Biochemical and structural analysis are the most common ways to elucidate the multiplex mechanisms of proteins. However, several studies such as protein crystallography and enzyme kinetics require a large amount of pure proteins for analysis. Thus, the Escherichia coli expression system providing a higher production level of recombinant proteins and various fusion tags for affinity purification meet the needs of the sample preparation. However, it is more difficult to express eukaryotic proteins in the prokaryotic system due to the protein folding usually required approximated post translational modifications or a specific chaperon system. With the maturity of genetic engineering technology, the plant-based expression systems expression systems has a great research potential to produce the eukaryotic recombinant proteins. Agrobacterium-mediated protein transient expression system uses Agrobacterium as a gene delivery vector to transfect target genes into epidermis of plant leaves and the target gene can be transiently expressed in 2 to 6 days. It is expected to be the solution overcoming the bottleneck of eukaryotic protein research in the past. Therefore, we used green fluorescence protein (GFP) as a model protein to establish the optimal procedure for transient protein expression and purification. Following the established procedure, the expression, purification and further enzymatic analysis of AtKAI2 were examined. We also try to overcome the bottleneck in obtaining plant membrane proteins. We successfully purified VrVPPase and performed single particle analysis of VrVPPase. Taken together, these results showed that agroinfiltration system is a high potential tool for producing plant recombinant proteins for investigating the molecular mechanisms and regulation in the plants.

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