Biosynthesis of (Bacterio) chlorophylls ATP-DEPENDENT TRANSIENT SUBUNIT INTERACTION AND ELECTRON TRANSFER OF DARK OPERATIVE PROTOCHLOROPHYLLIDE OXIDOREDUCTASE
Biosynthesis of (Bacterio) chlorophylls ATP-DEPENDENT TRANSIENT SUBUNIT INTERACTION AND ELECTRON TRANSFER OF DARK OPERATIVE PROTOCHLOROPHYLLIDE OXIDOREDUCTASE
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DOI：
10.1074/jbc.m109.087874
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发表时间：
2010-03-12
影响因子：
4.8
通讯作者：
Jahn, Dieter
中科院分区：
文献类型：
--
作者：
Broecker, Markus J.;Waetzlich, Denise;Jahn, Dieter
暗作用原叶绿素氧化还原酶（DPOR）催化原叶绿素a的不依赖光的双电子还原形成叶绿素a，这是叶绿素a和细菌叶绿素a生物合成的最后一个共同前体。在atp依赖性DPOR催化过程中，携带[4Fe-4S]簇的同二聚体ChlL(2)亚基将电子转移到相应的异四聚体催化亚基（ChlN/ChlB）(2)，该亚基也具有氧化还原活性[4Fe-4S]簇。为了研究这两种亚配合物的瞬态相互作用和由此产生的电子转移反应，在与不可水解的ATP类似物腺苷5‘（γ -硫）三磷酸、腺苷5’-（β， γ -氨基）三磷酸或MgADP与AlF4-结合孵育后，将蓝藻原绿球藻中含有亚基ChlN、ChlB和ChlL的三元dor酶全配合物捕获为八聚体（ChlN/ChlB）(2)(ChlL（2))(2)）配合物。另外，在开关II区缺失一个Leu（153）的突变体ChlL(2)蛋白也允许形成稳定的八聚体复合物。此外，有效的复合体形成需要原叶绿内酯的存在。三元DPOR配合物的电子顺磁共振波谱显示位于ChlL上的[4Fe-4S]团簇被还原(2)，表明ATP完全水解是亚基间电子转移的先决条件。圆二色光谱实验表明，ATP结合后ChlL(2)的构象发生了核苷酸依赖的变化。总结了一个核苷酸依赖的开关机制，触发三元配合物的形成和电子转移。根据这些结果，推导出了DPOR催化氧化还原的详细循环。
Dark operative protochlorophyllide oxidoreductase (DPOR) catalyzes the light-independent two-electron reduction of protochlorophyllide a to form chlorophyllide a, the last common precursor of chlorophyll a and bacteriochlorophyll a biosynthesis. During ATP-dependent DPOR catalysis the homodimeric ChlL(2) subunit carrying a [4Fe-4S] cluster transfers electrons to the corresponding heterotetrameric catalytic subunit (ChlN/ChlB)(2), which also possesses a redox active [4Fe-4S] cluster. To investigate the transient interaction of both subcomplexes and the resulting electron transfer reactions, the ternary DPOR enzyme holocomplex comprising subunits ChlN, ChlB, and ChlL from the cyanobacterium Prochlorococcus marinus was trapped as an octameric (ChlN/ChlB)(2)(ChlL(2))(2) complex after incubation with the nonhydrolyzable ATP analogs adenosine 5'(gamma-thio) triphosphate, adenosine 5'-(beta,gamma-imido) triphosphate, or MgADP in combination with AlF4-. Aitionally, a mutant ChlL(2) protein, with a deleted Leu(153) in the switch II region also allowed for the formation of a stable octameric complex. Furthermore, efficient complex formation required the presence of protochlorophyllide. Electron paramagnetic resonance spectroscopy of ternary DPOR complexes revealed a reduced [4Fe-4S] cluster located on ChlL(2), indicating that complete ATP hydrolysis is a prerequisite for intersubunit electron transfer. Circular dichroism spectroscopic experiments indicated nucleotide-dependent conformational changes for ChlL(2) after ATP binding. A nucleotide-dependent switch mechanism triggering ternary complex formation and electron transfer was concluded. From these results a detailed redox cycle for DPOR catalysis was deduced.