Díaz Adelaida, Valdés Víctor-Julián, Rudiño-Piñera Enrique, Horjales Eduardo, Hansberg Wilhelm
Instituto de Fisiología Celular, Universidad Nacional Autónoma de México, Circuito exterior s/n, Ciudad Universitaria, México, D. F. CP 04510, México.
J Mol Biol. 2009 Feb 13;386(1):218-32. doi: 10.1016/j.jmb.2008.12.019. Epub 2008 Dec 14.
Neurospora crassa has two large-subunit catalases, CAT-1 and CAT-3. CAT-1 is associated with non-growing cells and accumulates particularly in asexual spores; CAT-3 is associated with growing cells and is induced under different stress conditions. It is our interest to elucidate the structure-function relationships in large-subunit catalases. Here we have determined the CAT-3 crystal structure and compared it with the previously determined CAT-1 structure. Similar to CAT-1, CAT-3 hydrogen peroxide (H(2)O(2)) saturation kinetics exhibited two components, consistent with the existence of two active sites: one saturated in the millimolar range and the other in the molar range. In the CAT-1 structure, we found three interesting features related to its unusual kinetics: (a) a constriction in the channel that conveys H(2)O(2) to the active site; (b) a covalent bond between the tyrosine, which forms the fifth coordination bound to the iron of the heme, and a vicinal cysteine; (c) oxidation of the pyrrole ring III to form a cis-hydroxyl group in C5 and a cis-gamma-spirolactone in C6. The site of heme oxidation marks the starts of the central channel that communicates to the central cavity and the shortest way products can exit the active site. CAT-3 has a similar constriction in its major channel, which could function as a gating system regulated by the H(2)O(2) concentration before the gate. CAT-3 functional tyrosine is not covalently bonded, but has instead the electron relay mechanism described for the human catalase to divert electrons from it. Pyrrole ring III in CAT-3 is not oxidized as it is in other large-subunit catalases whose structure has been determined. Different in CAT-3 from these enzymes is an occupied central cavity. Results presented here indicate that CAT-3 and CAT-1 enzymes represent a functional group of catalases with distinctive structural characteristics that determine similar kinetics.
粗糙脉孢菌有两种大亚基过氧化氢酶，即CAT-1和CAT-3。CAT-1与非生长细胞相关，尤其在无性孢子中积累；CAT-3与生长细胞相关，并在不同应激条件下被诱导。我们感兴趣的是阐明大亚基过氧化氢酶的结构-功能关系。在此，我们确定了CAT-3的晶体结构，并将其与先前确定的CAT-1结构进行了比较。与CAT-1相似，CAT-3的过氧化氢（H₂O₂）饱和动力学表现出两个组分，这与存在两个活性位点一致：一个在毫摩尔范围内饱和，另一个在摩尔范围内饱和。在CAT-1结构中，我们发现了与其异常动力学相关的三个有趣特征：（a）将H₂O₂输送到活性位点的通道中有一个收缩处；（b）形成与血红素铁的第五个配位键的酪氨酸与相邻的半胱氨酸之间存在共价键；（c）吡咯环III氧化，在C5形成顺式羟基，在C6形成顺式γ-螺内酯。血红素氧化位点标志着通向中央腔的中央通道的起点，也是产物离开活性位点的最短路径。CAT-3在其主要通道中也有类似的收缩处，它可以作为一个由门之前的H₂O₂浓度调节的门控系统。CAT-3的功能性酪氨酸没有共价键结合，而是具有为人过氧化氢酶描述的电子中继机制，以从其上转移电子。CAT-3中的吡咯环III不像已确定结构的其他大亚基过氧化氢酶那样被氧化。与这些酶不同的是，CAT-3有一个被占据的中央腔。此处给出的结果表明，CAT-3和CAT-1酶代表了具有独特结构特征的过氧化氢酶功能组，这些特征决定了相似的动力学。
