Brandsch R, Bichler V
Biochemisches Institut der Universität, Freiburg, FRG.
FEBS Lett. 1987 Nov 16;224(1):121-4. doi: 10.1016/0014-5793(87)80433-7.
E. coli cells harbouring the recombinant plasmid pDB222 with the 6-HDNO gene under the control of the tac-promotor were induced with IPTG to synthesize a high amount of 6-HDNO protein. Part of this protein was present as 6-HDNO apoenzyme. The proportion of 6-HDNO apoenzyme formed could be increased when the induction of 6-HDNO synthesis by IPTG was performed in the presence of the inhibitor diphenyleneiodonium. The 6-HDNO apoenzyme thus formed could be transformed into enzymatically active holoenzyme in the presence of FAD by a process requiring an energy-generating system consisting of ATP, phosphoenolpyruvate and pyruvate kinase. This finding suggests that an enzymatic step(s) is (are) involved in the covalent flavinylation of 6-HDNO.
携带重组质粒pDB222的大肠杆菌细胞，其6 - HDNO基因受tac启动子控制，用异丙基 - β - D - 硫代半乳糖苷（IPTG）诱导以合成大量的6 - HDNO蛋白。该蛋白的一部分以6 - HDNO脱辅基酶形式存在。当在抑制剂二苯碘鎓存在下用IPTG诱导6 - HDNO合成时，形成的6 - HDNO脱辅基酶的比例会增加。如此形成的6 - HDNO脱辅基酶在黄素腺嘌呤二核苷酸（FAD）存在下，通过一个需要由三磷酸腺苷（ATP）、磷酸烯醇丙酮酸和丙酮酸激酶组成的能量产生系统的过程，可转化为具有酶活性的全酶。这一发现表明，在6 - HDNO的共价黄素化过程中涉及一个或多个酶促步骤。
