5-Aminolevulinate (ALA) synthase (ALAS), a homodimeric pyridoxal-5'-phosphate (PLP)-dependent enzyme, catalyzes the first step of heme biosynthesis in metazoa, fungi and α-proteobacteria. In this review, we focus on the advances made in unraveling the mechanism of the ALAS-catalyzed reaction during the past decade. The interplay between the PLP cofactor and the protein moiety determines and modulates the multi-intermediate reaction cycle of ALAS, which involves the decarboxylative condensation of two substrates, glycine and succinyl-CoA. Substrate binding and catalysis are rapid, and product (ALA) release dominates the overall ALAS kinetic mechanism. Interconversion between a catalytically incompetent, open conformation and a catalytically competent, closed conformation is linked to ALAS catalysis. Reversion to the open conformation, coincident with ALA dissociation, defines the slowest step of the reaction cycle. These findings were further substantiated by introducing seven mutations in the16-amino acid loop that gates the active site, yielding an ALAS variant with a greatly increased rate of catalytic turnover and heightened specificity constants for both substrates. Recently, molecular dynamics (MD) simulation analysis of various dimeric ALAS forms revealed that the seven active site loop mutations caused the proteins to adopt different conformations. In particular, the emergence of a β-strand in the mutated loop, which interacted with two preexisting β-strands to form an anti-parallel three-stranded β-sheet, conferred the murine heptavariant with a more stable open conformation and prompted faster product release than wild-type mALAS2. Moreover, the dynamics of the mALAS2 active site loop anti-correlated with that of the 35 amino acid C-terminal sequence. This led us to propose that this C-terminal extension, which is absent in prokaryotic ALASs, finely tunes mammalian ALAS activity. Based on the above results, we extend our previous proposal to include that discovery of a ligand inducing the mammalian C-terminal extension to fold offers a good prospect for the development of a new drug for X-linked protoporphyria and/or other porphyrias associated with enhanced ALAS activity and/or porphyrin accumulation.

中文翻译：

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5-氨基乙酰丙酸合酶催化：血红素生物合成中的捕手。

5-氨基乙酰丙酸盐（ALA）合酶（ALAS）是一种同二聚的吡ido醛5'-磷酸（PLP）依赖性酶，催化后生动物，真菌和α-变形杆菌中血红素生物合成的第一步。在这篇综述中，我们重点介绍了过去十年中在阐明ALAS催化反应机理方面取得的进展。PLP辅因子与蛋白质部分之间的相互作用决定并调节了ALAS的多中间体反应循环，该循环涉及两个底物甘氨酸和琥珀酰辅酶A的脱羧缩合。底物的结合和催化作用很快，产物（ALA）的释放主导了整个ALAS动力学机制。催化上不适合的开放构象和催化上合适的封闭构象之间的相互转化与ALAS催化有关。还原为开放构象，与ALA解离相吻合的是反应周期中最慢的一步。通过在门控活性位点的16个氨基酸的环中引入七个突变，进一步证实了这些发现，从而产生了ALAS变体，其催化转化率大大提高，并且两种底物的特异性常数均提高。最近，各种二聚ALAS形式的分子动力学（MD）模拟分析表明，七个活性位点环突变导致蛋白质采用不同的构象。特别是，在突变环中出现了一条β链，该链与两个预先存在的β链相互作用形成一个反平行的三链β折叠，使鼠类七变异体具有更稳定的开放构象并促进了更快的生成释放比野生型mALAS2。而且，mALAS2活性位点环的动力学与35个氨基酸的C端序列的反相关。这导致我们提出原核ALASs中不存在的C末端延伸可以很好地调节哺乳动物ALAS的活性。基于上述结果，我们将先前的提议扩展到包括诱导哺乳动物C端延伸折叠的配体的发现，为开发X连锁原卟啉和/或与之相关的其他卟啉症的新药物提供了良好的前景。增强ALAS活性和/或卟啉积聚。