First structure of full-length mammalian phenylalanine hydroxylase reveals the architecture of an autoinhibited tetramer
First structure of full-length mammalian phenylalanine hydroxylase reveals the architecture of an autoinhibited tetramer
复制标题
DOI：
10.1073/pnas.1516967113
复制
发表时间：
2016-03-01
影响因子：
11.1
通讯作者：
Jaffe, Eileen K.
中科院分区：
文献类型：
--
作者：
Arturo, Emilia C.;Gupta, Kushol;Jaffe, Eileen K.
提高对苯丙氨酸羟基酶(PAH)结构、动力学和功能之间关系的了解，可以导致治疗苯丙酮尿症所需的新疗法，苯丙酮尿症是氨基酸代谢最常见的先天错误。PAH是一种多结构域的同源多聚体蛋白，其构象和多聚化特性响应底物苯丙氨酸(Phe)的变构激活；变构调节是将Phe维持在神经毒性水平以下所必需的。最近提出的多环芳烃变构调节模型涉及主要的结构域运动和结构上不同的多环芳烃四聚体[Jaffe EK，Stith L，Lawrence SH，Andrake M，Dunbrack RL，Jr(2013)Arch Biochem BiPhys 530(2)：73-82]。在这里，我们提出了，据我们所知，第一个全长哺乳动物(大鼠)多环芳烃的自抑制构象的X射线晶体结构。在没有Phe的情况下，单分散四聚多环芳烃的层析分离有助于确定2.9埃的晶体结构。全长多环芳烃的结构取代了被广泛用于理性化苯丙酮尿症基因-表型关系的复合同源模型。小角X射线散射(SAXS)证实，在没有Phe的情况下，该四聚体占主导地位，不同于稳定的变构激活的PAH四聚体。缺乏激活的PAH的结构细节仍然是完全了解苯丙酮尿症的基因-表型关系的障碍。然而，结合使用SAXS和X射线结晶学来检查多环芳烃的结构，据我们所知，提供了第一个四聚体形式的酶的完整视图，这是以前的部分晶体结构所不可能的，并有助于解释到目前为止无法评估的丰富的生化和结构数据。
Improved understanding of the relationship among structure, dynamics, and function for the enzyme phenylalanine hydroxylase (PAH) can lead to needed new therapies for phenylketonuria, the most common inborn error of amino acid metabolism. PAH is a multidomain homo-multimeric protein whose conformation and multimerization properties respond to allosteric activation by the substrate phenylalanine (Phe); the allosteric regulation is necessary to maintain Phe below neurotoxic levels. A recently introduced model for allosteric regulation of PAH involves major domain motions and architecturally distinct PAH tetramers [Jaffe EK, Stith L, Lawrence SH, Andrake M, Dunbrack RL, Jr (2013) Arch Biochem Biophys 530(2): 73-82]. Herein, we present, to our knowledge, the first X-ray crystal structure for a full-length mammalian (rat) PAH in an autoinhibited conformation. Chromatographic isolation of a monodisperse tetrameric PAH, in the absence of Phe, facilitated determination of the 2.9 angstrom crystal structure. The structure of full-length PAH supersedes a composite homology model that had been used extensively to rationalize phenylketonuria genotype-phenotype relationships. Small-angle X-ray scattering (SAXS) confirms that this tetramer, which dominates in the absence of Phe, is different from a Phestabilized allosterically activated PAH tetramer. The lack of structural detail for activated PAH remains a barrier to complete understanding of phenylketonuria genotype-phenotype relationships. Nevertheless, the use of SAXS and X-ray crystallography together to inspect PAH structure provides, to our knowledge, the first complete view of the enzyme in a tetrameric form that was not possible with prior partial crystal structures, and facilitates interpretation of a wealth of biochemical and structural data that was hitherto impossible to evaluate.