植物的細胞壁主要由纖維素和半纖維素所組成，大約占所有組成的50%~60%，目前被認為是最主要的生質材料。科學家利用纖維素酶將纖維素和半纖維素這兩個主要的成分轉化為可發酵的醣類的方法已日益趨進重要，再加上這些可發酵的醣類是產生生物能源(bio-energy)最主要的原料，例如乙醇、丁醇、生質燃料等。在現今社會化石燃料日益短缺的問題下，再生能源的生產或許可以暫緩能源危機。目前在草食性動物瘤胃當中所居住的厭氧微生物由於草食性動物食性的關係，推測其可能具有多樣性纖維素分解酵素的活性，因此科學家認為這些微生物在分解纖維素上是相當重要的來源之一。在此研究當中，我們從努比亞山羊的瘤胃中發現一株具有纖維素酶活性的厭氧細菌。並且搭配上16S rDNA比對序列分析發現，這菌株是屬於鏈球菌屬(Streptococcus sp.)。鏈球菌是一種球狀的革蘭氏陽性菌，無法自行移動，並且不會產生孢子。在此研究中，將這菌株培養在以農業廢棄物作為碳源的培養液中，經由活性測試可以偵測到有較高的纖維素酶活性。這些纖維素水解酶的的活性，利用CMC-zymographic的方法更被為確定。接著依據已發表的Streptococcus gallolyticus的基因體序列去設計引子對(primer pair)，利用PCR，找到了1623bp，可以轉譯為540個胺基酸的纖維水解酵素，命名為Sg-EGV。這個纖維水解酵素屬於內切性的水解酶(endo-glucanase)，被歸類為水解酵素第五類。利用大腸桿菌去大量表現生產Sg-EGV，表現後的重組蛋白在溫度50℃、pH值為4的環境下可測得其最佳活性。Sg-EGV可以專一性的水解非結晶型木質纖維素的β-1,4的鍵結，例如:CMC (carboxymethylcellulose)或是大麥的β-D-glucan。在測試熱穩定的實驗中，將Sg-EGV放在60℃下，6小時，它的活性還保有原本的60%。在耐酸鹼的穩定上，蛋白可以穩定的存在介於pH 3到10之間。有一些研究指出，金屬二價離子可以提高纖維素酶的作用活性，Sg-EGV在有鈷和錳離子存在下，可以提高50%以上的活性。由厭氧環境中找到適合在酸性下作用的纖維水解蛋白，對於生質酒精的發展有一定的助益。

Cellulose and hemicelluloses comprise approximately 50 to 60% (w/w) of plant cell walls and are considered as the major components of the biomass. Bioconversion of these two components to fermentable sugars by celluases has become increasingly important, due to the potential for production of bio-energy, such as ethanol, butanol and other hydrocarbon biofuels. These so called renewable energies could significantly reduce fossil fuel consumption. Anaerobic microbes living in the rumen of herbivores possess diverse cellulase activities and represent potential sources of cellulases. In this study, a bacterium that exhibited strong cellulase activity on Congo red staining plate was isolated from the rumen of a Nubian Goat. Based on the 16S rRNA sequence, this bacterium was identified as Streptococcus sp. Streptococci are non-motile, gram-positive, non-sporeforming bacteria. By culturing this bacterium in the medium containing agricultural residues, high cellulase activity could be detected. The cellulase activity of crude protein was further confirmed by a CMC-zymographic analysis. Primer pair was then designed based on the Streptococcus gallolyticus genomic sequence to obtain endocellulase gene using PCR method. A full-length gene of 1623bp, Sg-EGV, encoding this glycosyl hydrolase of 540 amino acids was isolated. The endo-glucanase belongs to the glucoside hydrolase family V. The recombinant Sg-EGV protein expressed in Escherichia coli showed an optimal activity at 50℃, pH 4. Sg-EGV displayed high specific activity towards β-1,4-linkage of amorphous region of lignocelluloses such as carboxymethylcellulose and β-D-glucan from barley. Furthermore, the recombinant Sg-EGV retained more than 60% of its initial activity after 6 h of incubation at 60°C. Most importantly, the recombinant protein was stable in the pH range of 3-10. Several divalent cations were able to enhance the endo-glucanase activity; specifically, 10 mM CoCl2 and MnCl2 increase the enzyme activity more than 50%. This work identified an acidstable endo-glucanase, Sg-EGV from Streptococcus sp. that might be useful for cellulose hydrolysis for biofuel production.

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