Kankare J, Neal G S, Salminen T, Glumoff T, Glumhoff T [corrected to Glumoff T ], Cooperman B S, Lahti R, Goldman A
Centre for Biotechnology, Turku, Finland.
Protein Eng. 1994 Jul;7(7):823-30. doi: 10.1093/protein/7.7.823.
The structure of E.coli soluble inorganic pyrophosphatase has been refined at 2.7 A resolution to an R-factor of 20.9%. The overall fold of the molecule is essentially the same as yeast pyrophosphatase, except that yeast pyrophosphatase is longer at both the N- and C-termini. Escherichia coli pyrophosphatase is a mixed alpha + beta protein with a complicated topology. The active site cavity, which is also very similar to the yeast enzyme, is formed by seven beta-strands and an alpha-helix and has a rather asymmetric distribution of charged residues. Our structure-based alignment extends and improves upon earlier sequence alignment studies; it shows that probably no more than 14, not 15-17 charged and polar residues are part of the conserved enzyme mechanism of pyrophosphatases. Six of these conserved residues, at the bottom of the active site cavity, form a tight group centred on Asp70 and probably bind the two essential Mg2+ ions. The others, more spreadout and more positively charged, presumably bind substrate. Escherichia coli pyrophosphatase has an extra aspartate residue in the active site cavity, which may explain why the two enzymes bind divalent cation differently. Based on the structure, we have identified a sequence motif that seems to occur only in soluble inorganic pyrophosphatases.
大肠杆菌可溶性无机焦磷酸酶的结构已在2.7埃分辨率下精修，R因子为20.9%。该分子的整体折叠与酵母焦磷酸酶基本相同，只是酵母焦磷酸酶在N端和C端都更长。大肠杆菌焦磷酸酶是一种具有复杂拓扑结构的α+β混合蛋白。活性位点腔与酵母酶也非常相似，由七条β链和一条α螺旋形成，带电残基分布相当不对称。我们基于结构的比对扩展并改进了早期的序列比对研究；结果表明，参与焦磷酸酶保守酶机制的带电和极性残基可能不超过14个，而非15至17个。这些保守残基中有六个位于活性位点腔底部，以天冬氨酸70为中心形成紧密基团，可能结合两个必需的Mg2+离子。其他残基分布更分散且带正电荷更多，可能结合底物。大肠杆菌焦磷酸酶在活性位点腔中有一个额外的天冬氨酸残基，这可能解释了这两种酶结合二价阳离子方式不同的原因。基于该结构，我们确定了一个似乎仅存在于可溶性无机焦磷酸酶中的序列基序。
