鳥胺酸異構酶之特性分析

Studies on coenzyme B12-dependent D-Ornithine 4,5-aminomutase from Clostridium sticklandii

余琇婷(Hsiu-Ting Yu)

指導教授 ： 陳灝平

國立臺北科技大學/工程學院/生物科技研究所/碩士(2008年)

摘要

Clostridium sticklandii 此菌株內的鳥胺酸異構酶(D-Ornithine aminomutase)可將D-ornithine 催化成(2R，4R)-2，4-diaminopentanoic acid。鳥胺酸異構酶由兩個次單元所組成，分別為OraS及OraE。OraS由121個胺基酸組成，分子量為12800 Da；OraE 由753個胺基酸所組成，分子量為82900 Da。前人研究發現OraE及OraSE在部分金屬離子存在時，具有蛋白水解酶活性，本論文針對此蛋白酶活性，改建分析條件及測試酵素對各種金屬離子之動力學參數，並利用定點突變修飾胺基酸序列，尋找蛋白水解酶及金屬離子結合的活性部位。實驗結果顯示OraSE為金屬蛋白水解酶，並推測OraSE具有非專一性自我水解的特性。本論文亦發現鳥胺酸異構酶和同樣來自Clostridium sticklandii此菌株內的離胺酸異構酶（lysine 5,6-aminomutase），都對D-α-lysine具有催化能力並生成產物。經實驗分析測得OraSE對D-Lysine的K m值為5.0mM，OraSE對D-Lysine的催化活性明顯低於主要受質D-Ornithine。

關鍵字

鳥胺酸異構酶金屬蛋白水解酶D-Lysine

並列摘要

D-Ornithine aminomutase from Clostridium sticklandii catalyzes the 1,2-rearrangement reaction of D-Ornithine to (2R,4S)-2,4-diaminopentanoic acid. The enzyme comprises two subunits, OraS and OraE. Previous studies have shown that OraSE and OraE protein possess proteolytic activities in the presence of certain metal ions. The enzymatic assay method for this activity is established in this study. Site-directed mutagenesis studies of putative metal-binding are also carried out. My results confirm that OraSE belongs to a families of metalloprotease, and its substrate specificity is also investigated. Finally, I first show that the enzyme posseses novell activities toward D-a-Lysine. That Kcat and Km value for D-A_Lysine is also determind.

並列關鍵字

Ornithine aminomutase ； metalloprotease ； D-Lysine

參考文獻

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