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標題: | 斑節蝦眼的蛋白質牻牛兒牻牛兒基轉移?I之純化與定性 Purification and Characterization of Protein Geranylgeranyltransferase I from the Eyes of the Shrimp Penaeus japonicus |
作者: | 林睿軒 |
出版年 : | 1997 |
學位: | 碩士 |
摘要: | 本論文是以水生無脊椎動物斑節蝦的眼為材料,藉以勝?受質KLKCFFL的酵素活性檢測方法,先後經過HiTrap?Q陰離子交換管柱層析,Superdex TM200膠體過瀘管柱層析,Mono Q陰離子交換管柱層析,最後以勝?偶合親合力管柱完成蛋白質牻牛兒牻牛兒基轉移?I (PGGT I)的純化。純化的PGGT I比活性為188 units/mg。從膠體過濾管柱層析估算的分子量為67 kDa與SDS-PAGE所估算的66 kDa相近,推測斑節蝦蛋白質牻牛兒牻牛兒基轉移?I為單體結構。所得的蛋白質被抗法呢基轉移?α次單元的抗體(Y-53)所辨認,顯示二者含有相似的胺基酸序列。 在所檢測的勝?中,KCFFL是較好的勝?受質。酵素作用最適的酸鹼值是pH 8.0。斑節蝦蛋白質牻牛兒牻牛兒基轉移?I證實是一種金屬性?(EDTA的I.C.50=0.25μM)但活性會受Mg2+(I.C.50=500μM)及Zn2+ (I.C.50=50μM)所抑制。根據各組織的酵素活性檢測,廣布於斑節蝦的眼、肝胰臟、心臟、鰓、神經節、腸、肌肉及殼各組織。 Protein geranylgeranyltransferase I was purified from the eyes of shrimp Penaeus japonicus by the sequentail column chromatography on HiTrap? Q, Superdex? 200, Mono Q and peptide-coupled affinity column. The purified enzyme was found to have relative Mr of 66 kDa as estimated by SDS-PAGE. Since the active protein geranylgeranyltransferase I from Superdex? 200 was found to have relative mass of 67 kDa, the purified enzyme was deduced to be a monomer. Synthetic peptide KCFFL is the best substrate of the protein geranylgeranyltransferase I among the investigated peptides. The optimal pH for enzyme activity is pH 8.0. The enzyme was inhibited by Mg++ and Zn++ ions at 50μM and 500μM respectively. Since EDTA (IC50=0.25μM) is inhibitory as well, the protein geranylgeranyltransferase I of Penaeus japonicus is a metaloenzyme. |
URI: | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/76304 |
全文授權: | 未授權 |
顯示於系所單位: | 動物學研究所 |
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