The crystal structure of a bacterial Class II ketol-acid reductolsomerase: Domain conservation and evolution
The crystal structure of a bacterial Class II ketol-acid reductolsomerase: Domain conservation and evolution
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DOI：
10.1110/ps.051791305
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发表时间：
2005-12-01
期刊：
影响因子：
8
通讯作者：
Duggleby, RG
中科院分区：
文献类型：
--
作者：
Tyagi, R;Duquerroy, S;Duggleby, RG
酮醇酸还原异构酶（KARI; EC 1.1.1.86）催化支链氨基酸生物合成中的两个步骤。跨物种的氨基酸序列比较表明，有两种类型的这种酶：在真菌和大多数细菌中发现的短形式（1类）和植物中典型的长形式（11类）。每一种的晶体结构以前都有报道。然而，大肠杆菌等一些细菌具有较长的形式，其中的氨基酸序列与植物中发现的氨基酸序列明显不同。在这里，我们报道了E.大肠杆菌酶在2.6 A分辨率，第一个三维结构的任何细菌11类KARI。该酶由两个结构域组成，一个具有混合的α/β结构，这与其他吡啶核苷酸依赖性内切酶中发现的结构域相似。第二个结构域主要是α-螺旋，并显示出内部复制的强有力证据。比较了E.大肠杆菌、铜绿假单胞菌和菠菜中的氨基酸序列表明，大多数残基占据活性位点的保守位置。E. coli KARI结晶为四聚体，可能是生物活性单位。这与形成十二聚体的铜绿假单胞菌KARI和形成二聚体的菠菜KARI形成对比。在急诊coli KARI四聚体是一种新型的亚基间相互作用表面，由一对对称的球状突起形成。
Ketol-acid reductoisomerase (KARI; EC 1.1.1.86) catalyzes two steps in the biosynthesis of branched-chain amino acids. Amino acid sequence comparisons across species reveal that there are two types of this enzyme: a short form (Class 1) found in fungi and most bacteria, and a long form (Class 11) typical of plants. Crystal structures of each have been reported previously. However, some bacteria such as Escherichia coli possess a long form, where the amino acid sequence differs appreciably from that found in plants. Here, we report the crystal structure of the E. coli enzyme at 2.6 A resolution, the first three-dimensional structure of any bacterial Class 11 KARI. The enzyme consists of two domains, one with mixed alpha/beta structure, which is similar to that found in other pyridine nucleotide-dependent dehydrogenases. The second domain is mainly alpha-helical and shows strong evidence of internal duplication. Comparison of the active sites between KARI of E. coli, Pseudomonas aeruginosa, and spinach shows that most residues occupy conserved positions in the active site. E. coli KARI was crystallized as a tetramer, the likely biologically active unit. This contrasts with P. aeruginosa KARI, which forms a dodecamer, and spinach KARI, a dimer. In the E. coli KARI tetramer, a novel subunit-to-subunit interacting surface is formed by a symmetrical pair of bulbous protrusions.