離胺酸異構酶性質之探討

Characterization of Lysine 5,6-aminomutase from Clostridium sticklandii

黃品慈(Ping-Tzu Huang)

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此菌株內的離胺酸異構酶（lysine 5,6-aminomutase）可將D-α-lysine催化成2,5-diaminohexanoic acid。離胺酸異構酶目前已知二個次單元kamD與kamE，KamD由516個胺基酸所組成，分子量為51,000 Da；KamE由262個胺基酸所組成，分子量為30,000 Da。KamDE在催化的過程中，需要輔酶B6 ( PLP )及輔酶B12 ( AdoCbl )兩個輔酶參與反應。本篇論文證實KamDE與AdoCbl、methylcobalamin ( MeCbl )及methylcobinamide ( Mecbi )會和KamDE形成base-off / histidine on之結構，而adenosylcobinamide ( Adocbi )為base-off / histidine on及base-off / histidine off之混合型結構。目前研究指出KamDE與鳥胺酸異構酶S次單元（OraS）能夠形成複合體。OraS的存在會降低AdoCbl、MeCbl、Mecbi及AdoCbi對KamDE之結合力。另外，AdoCbi取代AdoCbl與KamDE反應時，OraS失去異位活化的能力。另一個B12類似物adenosylcobinamide-GDP ( Adocbi-GDP )無法與KamDE結合在一起。

關鍵字

輔酶B12 ； 離胺酸異構酶鳥胺酸異構酶

並列摘要

Lysine 5,6-aminomutase from Clostridium sticklandii catalyzes the interconversion of D-α-lystine into 2,5-diaminohexanoic acid. The enzyme comprises two different subunits, KamD (Mr 51,000) and KamE (Mr 30,000). Two different coenzymes, pyridoxal phosphate (PLP) and adenosylcobalamin (AdoCbl) , are involved in this enzymatic reaction. In this study, my results show that KamDE binds AdoCbl, adenosylcobinamide (AdoCbi), methylcobinamide (MeCbi), and methylcobalamin (MeCbl) in the “base-off / histidine on” mode. However, the other cofactor, adenosylcobinamide-GDP (AdoCbi-GDP), can not be bound by the enzyme. The presence of OraS raises the Kd of KamDE for AdoCbl、MeCbi and AdoCbi and the Km of KamDE for AdoCbi, suggesting that OraS reduces the apparent affinity for the cofactor.

並列關鍵字

AdoCbl ； lysine aminomutase ； ornithine aminomutase

參考文獻

[17] 曾振興，離胺酸異構酶與鳥胺酸異構酶S次單元之交互作用，碩士論文，國立臺北科技大學生物科技研究所，台北，2007。

[1] E. N. G. Marsh, and D. P. Ballou, "Coupling of cobalt-carbon bond homolysis and hydrogen atom abstraction in adenosylcobalamin-dependent glutamate mutase ," Biochemistry, vol. 37, no. 34, 1998, pp. 11864-11872.

[2] J. K. Dyer, and R. N. Costilow, "2,4-diaminovaleric acid: an intermediate in the anaerobic oxidation of ornithine by Clostridium sticklandii," J. Bacteriol. vol. 101, no. 1, 1970, pp. 77-83.

[4] H. P. Chen, F. D. Lung, C. C. Yeh, H. L. Chen, and S. H. Wu, "The role of the conserved histidine-aspartate pair in the “base-off” binding of cobalamins," Bioorg. Med. Chem. vol. 12, no. 3, 2004, pp. 577-582.

[5] C. H. Chang, and P. A. Fery, "Cloning, sequencing, heterologous expression, purification, and characterization of adenosylcobalamin-dependent D-lysine 5,6-aminomutase from Clostridium stickland," J. Biol. Chem. vol. 275, no. 1, 2000, pp. 106-114.