培養基中金屬離子對枯草桿菌表面素產率之影響

The effect of metal ions on surfactin production by Bacillus subtilis

陳孟秋(Meng-Chiu Chen)

指導教授 ： 簡志青

共同指導教授 ： 康世旭

元智大學/工程學院/生物科技與工程研究所/碩士(2008年)

摘要

表面素(surfactin)是由Bacillus subtilis ATCC 21332 以非核醣體胜肽合成機制所合成的脂胜肽生物界面活性劑，結構包含七個氨基酸所組成的環狀胜肽鏈，及一長碳鏈的脂肪酸支鏈。表面素是目前文獻上發現最有效的生物界面活性劑。在應用上，因侷限於產量低、生產成本高的瓶頸，因此尚無法達到商業化之目的。因為 B. subtilis ATCC 21332 的遺傳性質尚未確立且難以接受外來DNA，因此本研究將嘗試使用改質B. subtilis 168 所獲得的的高表面素產率菌株(B. subtlis SWK569)(2006，劉惠娟)研究與表面素產率相關的可能基因。研究發現MS培養基中添加1 mM 的Fe2+ 以及10 µM Mn2+，可以使SWK569在培養基累積表面素的濃度至5223 mg/L，為只含有2mM Fe2+的培養基的10 倍。這個研究除了可實際應用於表面素生產外，也有助於後續對表面素生產調控機制的研究。由SDS-PAGE 顯示，可發現添加Fe2+以及Mn2+會影響SWK569 的蛋白質表現，經過分析比對資料庫後，發現有許多蛋白質需要二價金屬離子作為輔助因子(cofactor)，在這些蛋白質中亦發現有部分和表面素合成有直接相關相關的蛋白質。

關鍵字

枯草桿菌 ； 表面素

並列摘要

Surfactin is synthesized by Bacillus subtilis through a non-ribosomal peptide biosynthesis mechanism, and it consists of seven amino acid peptides, arranged in a lactone ring and bonded to a β-hydroxy fatty acid. However, surfactin is not widely used because of the high production costs, which primarily due to low strain yields and high recovery expenses. Because of the B. subtilis ATCC 21332 is hard to accept external DNA, therefore this research will attempt uses this laboratory to change nature B. subtilis 168 - SWK series which have high surfactin productivity will be the main research. When we changed the concentration of Fe2+ to 1mM and Mn2+ to 10µM in the MS medium, we found that the concentration of surfactin in the medium was 5223 mg/L, and it was ten times higher than the original medium. This research
besides may the practical application to the surfactin production, also helpful to study the regulative mechanism research for surfactin production. The result of SDS-PAGE displays that Fe2+ and Mn2+ were the effect protein expression.

並列關鍵字

Bacilllus subtilis ； surfactin

參考文獻

魏毓宏 (2001). "二價金屬無機鹽類之添加對生物界面活性劑表面素生產之影響." 化工 48(3).

Al-Ajlani, M. M., Sheikh, M.A., Ahmad, Z., Hasnain, S. (2007). "Production of surfactin from Bacillus subtilis MZ-7 grown on pharmamedia commercial medium." Microbial Cell Factories 6.

Arfin, S. M., Umbarger, H.E. (1969). "Purification and properties of the acetohydroxy acid isomeroreductase of Salmonella typhimurium." Journal of Biological Chemistry 244(5): 1118-1127.

Arima, K., Kakinuma, A., Tamura, G. (1968). "Surfactin, a crystalline peptidelipid surfactant produced by Bacillus subtilis: isolation, characterization and its inhibition of fibrin clot formation." Biochemical and Biophysical Research Communications 31(3): 488-494.

Asselineau, C., Asselineau, J. (1978). "Trehalose-containing glycolipids." Progress in the chemistry of fats and other lipids 16: 59-99.