Structure/function studies on a S-adenosyl-L-methionine-dependent uroporphyrinogen III C methyltransferase (SUMT), a key regulatory enzyme of tetrapyrrole biosynthesis.
Structure/function studies on a S-adenosyl-L-methionine-dependent uroporphyrinogen III C methyltransferase (SUMT), a key regulatory enzyme of tetrapyrrole biosynthesis.
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DOI：
10.1016/j.jmb.2004.09.020
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发表时间：
2004-11
影响因子：
5.6
通讯作者：
J. Vévodová;R. Graham;E. Raux;H. Schubert;D. Roper;A. Brindley;A. Ian Scott;C. Roessner;N. Stamford;M. Elizabeth Stroupe;E. Getzoff;M. Warren;K. Wilson
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文献类型：
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作者：
J. Vévodová;R. Graham;E. Raux;H. Schubert;D. Roper;A. Brindley;A. Ian Scott;C. Roessner;N. Stamford;M. Elizabeth Stroupe;E. Getzoff;M. Warren;K. Wilson
由cobA基因编码的反硝化假单胞菌s -腺苷基-l-蛋氨酸依赖的尿卟啉原III甲基转移酶（SUMT）的晶体结构已通过分子置换达到2.7Å分辨率。SUMT是一种分支点酶，通过控制维生素b12和西罗血红素等化合物的通量，并催化尿卟啉原III转化为前体蛋白-2，在修饰四吡啶的生物合成中起关键作用。该酶的整体拓扑结构与sirohaem合成酶（CysG）和钴预corcorin -4甲基转移酶cif的SUMT模块相似，并且与后者结构一样，SUMT的产物s -腺苷-l-同型半胱氨酸结合在晶体中。讨论了SUMT结构中一些残基的作用，以及它们在更广泛的钴胺素生物合成甲基转移酶家族或SUMT成员亚群中的保护作用。通过诱变cobA基因产生SUMT的D47N、L49A、F106A、T130A、Y183A和M184A变异体，并进行SAM结合和酶活性检测。在这些变异中，只有D47N和L49A结合了共底物s -腺苷-l-蛋氨酸。因此，所有突变体合成预corrin-2的能力都受到严重限制，尽管D47N和L49A突变体都产生了大量的预corrin-1，这是一种europorphyrin原III的单甲基化衍生物。这些变异的活性是根据酶的结构来解释的。
The crystallographic structure of the Pseudomonas denitrificans S-adenosyl-l-methionine-dependent uroporphyrinogen III methyltransferase (SUMT), which is encoded by the cobA gene, has been solved by molecular replacement to 2.7Å resolution. SUMT is a branchpoint enzyme that plays a key role in the biosynthesis of modified tetrapyrroles by controlling flux to compounds such as vitamin B12and sirohaem, and catalysing the transformation of uroporphyrinogen III into precorrin-2. The overall topology of the enzyme is similar to that of the SUMT module of sirohaem synthase (CysG) and the cobalt-precorrin-4 methyltransferase CbiF and, as with the latter structures, SUMT has the product S-adenosyl-l-homocysteine bound in the crystal. The roles of a number of residues within the SUMT structure are discussed with respect to their conservation either across the broader family of cobalamin biosynthetic methyltransferases or within the sub-group of SUMT members. The D47N, L49A, F106A, T130A, Y183A and M184A variants of SUMT were generated by mutagenesis of the cobA gene, and tested for SAM binding and enzymatic activity. Of these variants, only D47N and L49A bound the co-substrate S-adenosyl-l-methionine. Consequently, all the mutants were severely restricted in their capacity to synthesise precorrin-2, although both the D47N and L49A variants produced significant quantities of precorrin-1, the monomethylated derivative of uroporphyrinogen III. The activity of these variants is interpreted with respect to the structure of the enzyme.