Structure-Function Studies on the Sulfate Activating Enzymes

[904003] segel, Fisher，本研究探讨了“硫酸盐活化酶”、atp硫化酶和APS激酶的催化和调控特性的结构特征。这些酶按顺序催化无机硫酸盐同化过程中的前两个反应。研究了黄曲青霉的同六聚体ATP硫化酶，该酶被PAPS变构抑制。这种反馈过程是丝状真菌所特有的，很可能反映了PAPS在真菌中是一个主要的分支点代谢物，而在其他生物中则不是。真菌ATP硫酰化酶的变构结构域位于多肽链的c端，与apskin酶具有高度的同源性。实验旨在确定(a)变构结构域的三维结构与真正的APS激酶的三维结构之间的关系，以及(b)由变构效应物诱导的过渡的结构性质。P. chrysogenum APS激酶，一种同型二聚体，也被研究以建立以下结构基础：(a)对磺核苷酸的高特异性，(b)底物的强制有序结合（是动力学的还是基于结构的？）以及(c)亚基结合与活性位点形成之间的关系。硫是构成活细胞的六种主要元素之一。含有硫的关键生物分子包括蛋白质氨基酸、半胱氨酸和蛋氨酸，以及维生素泛酸、生物素和维生素b1。地球表面（土壤和海洋等）的大多数硫以无机硫酸盐的形式存在。本项目重点研究ATP硫化酶和APS激酶的结构和功能，这两种酶催化无机硫酸盐进入生物分子的前两个反应。这些酶将从丝状真菌青霉菌中纯化出来（青霉菌能产生另一种重要的含硫化合物青霉素）。

9904003Segel, Fisher This research examines the structural features underlying the catalyticand regulatory properties of the "sulfate activating enzymes", ATPsulfurylase and APS kinase. These enzymes catalyze, in order, the first tworeactions in the assimilation of inorganic sulfate. The investigation focuses on the homo-hexameric ATP sulfurylase of Penicillium chrysogenum,which is allosterically inhibited by PAPS. This feedback process is uniqueto filamentous fungi, very likely reflecting the fact that PAPS is a majorbranch point metabolite in fungi, but not in other organisms. Theallosteric domain of fungal ATP sulfurylase is located in the C-terminalregion of the polypeptide chain and has a high degree of homology with APSkinase. Experiments are designed to determine (a) the relationshipbetween the three dimensional structure of the allosteric domain and thatof true APS kinase and (b) the structural nature of the transition inducedby the allosteric effector. P. chrysogenum APS kinase, a homodimer, isalso be studied in order to establish the structural basis of (a) the highspecificity for sulfonucleotides, (b) the compulsory ordered binding ofsubstrates (is it kinetic or structurally-based?), and (c) the relationshipbetween subunit association and formation of the active site.Sulfur is one of the six major elements that compose living cells. Keybiological molecules that contain sulfur include the protein amino acids,cysteine and methionine, and the vitamins pantothenic acid, biotin, andthiamin. Most of the sulfur on the Earth's surface (in the soil and oceans,etc.) is present as inorganic sulfate. This project focuses on the structures and functions of ATP sulfurylase and APS kinase, enzymes that catalyze the first two reactions in the incorporation of inorganic sulfate into biomolecules. The enzymes will be purified from the filamentous fungus, Penicillium chrysogenum (which produces another important sulfur compound, penicillin.)