Arginase (EC 3.5.3.1) catalyzes the hydrolysis of L-arginine to L-ornithine and urea, and requires a bivalent cation, especially Mn²⁺ for its catalytic activity. It is a component of the urea cycle and regulates the intracellular levels of l-arginine, which makes the arginase a target for treatment of vascular diseases and asthma. Mammalian arginases contain an unusual S-shaped motif located at the intermonomeric interface. Until now, the studies were limited to structural role of the motif. Then, our interest was focused on functional aspects and our hypothesis has been that the motif is essential for maintain the oligomeric state, having Arg308 as a central axis. Previously, we have shown that the R308A mutant is monomeric and re-associates to the trimeric-cooperative state in the presence of low concentrations of guanidine chloride.

We have now mutated Asp204 that interacts with Arg308 in the neighbor subunit, and also we mutated Glu256, proposed as important for oligomerization. Concretely, the human arginase I mutants D204A, D204E, E256A, E256Q and E256D were generated and examined. No differences were observed in the kinetic parameters at pH 9.5 or in tryptophan fluorescence. However, the D204A and E256Q variants were monomeric. On the other hand, D204E and E256D proved to be trimeric and kinetically cooperative at pH 7.5, whereas hyperbolic kinetics was exhibited by E256A, also trimeric.

The results obtained strongly support the importance of the interaction between Arg255 and Glu256 in the cooperative properties of arginase, and Asp204 would be relevant to maintain the oligomeric state through salt bridges with Arg255 and Arg308.

精氨酸酶 (EC 3.5.3.1) 催化 L-精氨酸水解为 L-鸟氨酸和尿素，并且需要二价阳离子，尤其是 Mn ²⁺才能发挥其催化活性。它是尿素循环的一个组成部分，调节细胞内l-精氨酸，使精氨酸酶成为治疗血管疾病和哮喘的靶点。哺乳动物精氨酸酶包含一个不寻常的 S 形基序，位于单体间界面。到目前为止，这些研究仅限于主题的结构作用。然后，我们的兴趣集中在功能方面，我们的假设是该基序对于维持寡聚状态必不可少，以 Arg308 作为中心轴。以前，我们已经表明 R308A 突变体是单体的，并在低浓度的氯化胍存在下重新关联到三聚体合作状态。

我们现在已经突变了与相邻亚基中的 Arg308 相互作用的 Asp204，并且我们也突变了 Glu256，它被认为对寡聚化很重要。具体而言，产生并检查了人精氨酸酶 I 突变体 D204A、D204E、E256A、E256Q 和 E256D。在 pH 9.5 或色氨酸荧光中没有观察到动力学参数的差异。然而，D204A 和 E256Q 变体是单体。另一方面，D204E 和 E256D 在 pH 7.5 下被证明是三聚体和动力学协同，而 E256A 表现出双曲线动力学，也是三聚体。

获得的结果有力地支持了 Arg255 和 Glu256 之间相互作用在精氨酸酶协同特性中的重要性，并且 Asp204 将与通过与 Arg255 和 Arg308 的盐桥保持寡聚状态有关。