Sakuno Emi, Wen Ying, Hatabayashi Hidemi, Arai Hatsue, Aoki Chiemi, Yabe Kimiko, Nakajima Hiromitsu
National Food Research Institute, Tsukuba, Ibaraki 305-8642, Japan.
Appl Environ Microbiol. 2005 Jun;71(6):2999-3006. doi: 10.1128/AEM.71.6.2999-3006.2005.
In the aflatoxin biosynthetic pathway, 5'-oxoaverantin (OAVN) cyclase, the cytosolic enzyme, catalyzes the reaction from OAVN to (2'S,5'S)-averufin (AVR) (E. Sakuno, K. Yabe, and H. Nakajima, Appl. Environ. Microbiol. 69:6418-6426, 2003). Interestingly, the N-terminal 25-amino-acid sequence of OAVN cyclase completely matched an internal sequence of the versiconal (VHOH) cyclase that was deduced from its gene (vbs). The purified OAVN cyclase also catalyzed the reaction from VHOH to versicolorin B (VB). In a competition experiment using the cytosol fraction of Aspergillus parasiticus, a high concentration of VHOH inhibited the enzyme reaction from OAVN to AVR, and instead VB was newly formed. The recombinant Vbs protein, which was expressed in Pichia pastoris, showed OAVN cyclase activity, as well as VHOH cyclase activity. A mutant of A. parasiticus SYS-4 (= NRRL 2999) with vbs deleted accumulated large amounts of OAVN, 5'-hydroxyaverantin, averantin, AVR, and averufanin in the mycelium. These results indicated that the cyclase encoded by the vbs gene is also involved in the reaction from OAVN to AVR in aflatoxin biosynthesis. Small amounts of VHOH, VB, and aflatoxins also accumulated in the same mutant, and this accumulation may have been due to an unknown enzyme(s) not involved in aflatoxin biosynthesis. This is the first report of one enzyme catalyzing two different reactions in a pathway of secondary metabolism.
在黄曲霉毒素生物合成途径中，胞质酶5'-氧杂averantin（OAVN）环化酶催化从OAVN到（2'S,5'S）-averufin（AVR）的反应（E. Sakuno、K. Yabe和H. Nakajima，《应用与环境微生物学》69:6418 - 6426，2003年）。有趣的是，OAVN环化酶的N端25个氨基酸序列与从其基因（vbs）推导的versiconal（VHOH）环化酶的内部序列完全匹配。纯化的OAVN环化酶也催化从VHOH到versicolorin B（VB）的反应。在使用寄生曲霉胞质部分的竞争实验中，高浓度的VHOH抑制了从OAVN到AVR的酶反应，取而代之的是新形成了VB。在毕赤酵母中表达的重组Vbs蛋白表现出OAVN环化酶活性以及VHOH环化酶活性。vbs缺失的寄生曲霉SYS - 4（= NRRL 2999）突变体在菌丝体中积累了大量的OAVN、5'-羟基averantin、averantin、AVR和averufanin。这些结果表明，由vbs基因编码的环化酶也参与黄曲霉毒素生物合成中从OAVN到AVR的反应。在同一突变体中也积累了少量的VHOH、VB和黄曲霉毒素，这种积累可能是由于与黄曲霉毒素生物合成无关的未知酶所致。这是关于一种酶在次生代谢途径中催化两种不同反应的首次报道。
