Ilari Andrea, Franceschini Stefano, Bonamore Alessandra, Arenghi Fabio, Botta Bruno, Macone Alberto, Pasquo Alessandra, Bellucci Luca, Boffi Alberto
Dipartimento di Scienze Biochimiche and Istituto di Biologia e Patologia Molecolari, CNR, Sapienza University, Piazzale Aldo Moro 5, 00185 Roma, Italy.
J Biol Chem. 2009 Jan 9;284(2):897-904. doi: 10.1074/jbc.M803738200. Epub 2008 Nov 12.
The enzyme norcoclaurine synthase (NCS) catalyzes the stereospecific Pictet-Spengler cyclization between dopamine and 4-hydroxyphenylacetaldehyde, the key step in the benzylisoquinoline alkaloid biosynthetic pathway. The crystallographic structure of norcoclaurine synthase from Thalictrum flavum in its complex with dopamine substrate and the nonreactive substrate analogue 4-hydroxybenzaldehyde has been solved at 2.1A resolution. NCS shares no common features with the functionally correlated "Pictet-Spenglerases" that catalyze the first step of the indole alkaloids pathways and conforms to the overall fold of the Bet v1-like protein. The active site of NCS is located within a 20-A-long catalytic tunnel and is shaped by the side chains of a tyrosine, a lysine, an aspartic, and a glutamic acid. The geometry of the amino acid side chains with respect to the substrates reveals the structural determinants that govern the mechanism of the stereoselective Pictet-Spengler cyclization, thus establishing an excellent foundation for the understanding of the finer details of the catalytic process. Site-directed mutations of the relevant residues confirm the assignment based on crystallographic findings.
去甲乌药碱合酶（NCS）催化多巴胺与4-羟基苯乙醛之间的立体特异性Pictet-Spengler环化反应，这是苄基异喹啉生物碱生物合成途径中的关键步骤。已解析出黄花唐松草去甲乌药碱合酶与多巴胺底物及非反应性底物类似物4-羟基苯甲醛形成的复合物的晶体结构，分辨率为2.1埃。NCS与催化吲哚生物碱途径第一步的功能相关“Pictet-Spengler酶”没有共同特征，并且符合类Bet v1蛋白的整体折叠结构。NCS的活性位点位于一条20埃长的催化隧道内，由一个酪氨酸、一个赖氨酸、一个天冬氨酸和一个谷氨酸的侧链构成其形状。氨基酸侧链相对于底物的几何结构揭示了控制立体选择性Pictet-Spengler环化反应机制的结构决定因素，从而为理解催化过程的更精细细节奠定了良好基础。对相关残基进行的定点突变证实了基于晶体学研究结果的归属。
