MacA is a second cytochrome c peroxidase of Geobacter sulfurreducens.
MacA is a second cytochrome c peroxidase of Geobacter sulfurreducens.
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MacA 是硫还原地杆菌的第二种细胞色素 c 过氧化物酶。
DOI：
10.1021/bi300249u
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发表时间：
2012
期刊：
影响因子：
2.9
通讯作者：
Einsle,Oliver
中科院分区：
文献类型：
--
作者：
Seidel,Julian;Hoffmann,Maren;Ellis,KatieE;Seidel,Antonia;Spatzal,Thomas;Gerhardt,Stefan;Elliott,SeanJ;Einsle,Oliver
金属还原δ-蛋白细菌硫还原地杆菌产生大量的OFC类细胞色素，其中许多参与了电子向不溶性金属氧化物的转移。其中，双性猕猴被指定为核心角色。在这里我们产生了G。硫磺还原蛋白MacA在大肠杆菌中的重组表达，解决了其在三种不同氧化状态下的三维结构。序列比较表明，该蛋白与ABTS-2为电子供体，具有过氧化氢还原酶活性。测得KM为38.5±3.7Mmin-1·mg-1，Vmax为0.78±0.03Mol·μ-1·mg-1，转化率为kCAT=0.46·S-1，未见Fe(III)还原酶活力。研究发现，MACA的电化学性质与其他细菌双氢过氧化物酶相似，此外，它还具有将电子传递给可溶性细胞色素PPCA的能力。CCPA和MACA之间的活性差异可以通过三个环区之一的结构变化来解释，环2在酶的还原激活过程中经历构象变化。这个环与活性部位的血红素相邻，形成一个开环结构，而不是像在CCPA中那样形成一个更刚性的螺旋。为了激活蛋白质，环必须将远端配体置换到环1中的活性部位H93。H93G变体在环2中意外地形成螺旋，在环1中无序，而改变电子转移血红素性质的M297H变体取消了还原激活。
The metal-reducing δ-proteobacteriumGeobacter sulfurreducensproduces a large number ofc-type cytochromes, many of which have been implicated in the transfer of electrons to insoluble metal oxides. Among these, the dihemic MacA was assigned a central role. Here we have producedG. sulfurreducensMacA by recombinant expression inEscherichia coliand have solved its three-dimensional structure in three different oxidation states. Sequence comparisons group MacA into the family of diheme cytochromecperoxidases, and the protein indeed showed hydrogen peroxide reductase activity with ABTS–2as an electron donor. The observedKMwas 38.5 ± 3.7 μM H2O2andvmaxwas 0.78 ± 0.03 μmol of H2O2·min–1·mg–1, resulting in a turnover numberkcat= 0.46 · s–1. In contrast, no Fe(III) reductase activity was observed. MacA was found to display electrochemical properties similar to other bacterial diheme peroxidases, in addition to the ability to electrochemically mediate electron transfer to the soluble cytochrome PpcA. Differences in activity between CcpA and MacA can be rationalized with structural variations in one of the three loop regions, loop 2, that undergoes conformational changes during reductive activation of the enzyme. This loop is adjacent to the active site heme and forms an open loop structure rather than a more rigid helix as in CcpA. For the activation of the protein, the loop has to displace the distal ligand to the active site heme, H93, in loop 1. A H93G variant showed an unexpected formation of a helix in loop 2 and disorder in loop 1, while a M297H variant that altered the properties of the electron transfer heme abolished reductive activation.