Mechanism and Stereochemistry of Enzymatic Cyclization of 24,30-Bisnor-2,3-oxidosqualene by Recombinant β-Amyrin Synthase

• Published in: Journal of the American Chemical Society 2004-06, Vol.126 (22), p.6880-6881
• Main Authors: Abe, Ikuro, Sakano, Yuichi, Sodeyama, Megumi, Tanaka, Hideya, Noguchi, Hiroshi, Shibuya, Masaaki, Ebizuka, Yutaka
• Format: Article
• Language: English
• Subjects: Biological and medical sciences; Chemistry; Cyclization; Exact sciences and technology; Fundamental and applied biological sciences. Psychology; Intramolecular Transferases - metabolism; Kinetics and mechanisms; Magnetic Resonance Spectroscopy; Mechanisms. Catalysis. Electron transfer. Models; Molecular biophysics; Molecular Structure; Organic chemistry; Physical chemistry in biology; Reactivity and mechanisms; Squalene - analogs & derivatives; Squalene - chemistry; Squalene - metabolism; Stereoisomerism
• ISSN: 0002-7863; 1520-5126
• DOI: 10.1021/ja0490368

Recombinant β-amyrin synthase from Pisum sativum converted 24,30-bisnor-2,3-oxidosqualene into a 3:1:0.2 mixture of 29,30-bisnor-β-amyrin, 29,30-bisnorgermanicol, and 29,30-bisnor-δ-amyrin. Further, enzyme reactions with [23-13C]- and [23,23-2H]-labeled isotopomers demonstrated that the cyclization did not proceed through formation of a lupanyl primary cation with a five-membered E-ring, but an electrophilic addition of the tetracyclic C-18 cation on to the terminal double bond directly generated a thermodynamically favored pentacyclic secondary cation with a less-strained six-membered E-ring. Interestingly, the formation of the three regioisomers suggested that the absence of the terminal methyl groups resulted in a structural perturbation in the folding conformation of the E-ring of the oleanyl cation at the active site of the enzyme.