Modification of vertebrate and algal prolyl 4-hydroxylases and vertebrate lysyl hydroxylase by diethyl pyrocarbonate. Evidence for histidine residues in the catalytic site of 2-oxoglutarate-coupled dioxygenases.
Modification of vertebrate and algal prolyl 4-hydroxylases and vertebrate lysyl hydroxylase by diethyl pyrocarbonate. Evidence for histidine residues in the catalytic site of 2-oxoglutarate-coupled dioxygenases.
复制标题
焦碳酸二乙酯对脊椎动物和藻类脯氨酰 4-羟化酶和脊椎动物赖氨酰羟化酶的修饰。
作者：
Raili Myllylä;V. Günzler;K. I. Kivirikko;D. Kaska
在2-酮戊二酸偶联双加氧酶的cDNA衍生的氨基酸序列中搜索保守的氨基酸残基，发现在这些蛋白质的C-末端区域存在两个不同的基序，间隔49-71个氨基酸。两个共同的基序中的每一个在保守位置处含有不变的组氨酸残基。2-酮戊二酸偶联双加氧酶在不同的过程中发挥作用，包括脊椎动物胶原蛋白中脯氨酸和赖氨酸残基的翻译后羟基化以及微生物头孢菌素的生物合成，但它们具有共同的反应机制，即需要在催化位点结合Fe 2+、2-酮戊二酸、O2和抗坏血酸。这两个区域的同源性，特别是相同的组氨酸，可能代表功能上重要的网站有关的催化活性。组氨酸残基的修饰由焦碳酸二乙酯灭活脊椎动物和藻类脯氨酰4-羟化酶和脊椎动物赖氨酰羟化酶，表明组氨酸残基在这些2-酮戊二酸偶联双加氧酶的催化位点中起作用。通过存在共底物而不是肽底物来防止失活。推测保守基序中的组氨酸残基可能具有Fe（2+）结合配体的功能。
A search for conserved amino acid residues within the cDNA-derived amino acid sequences of 2-oxoglutarate-coupled dioxygenases revealed the presence of two distinct motifs, spaced 49-71 amino acids apart, toward the C-terminal regions of these proteins. Each of the two common motifs contains an invariant histidine residue at a conserved position. The 2-oxoglutarate-coupled dioxygenases function in diverse processes, including the post-translational hydroxylation of proline and lysine residues in vertebrate collagens and the biosynthesis of microbial cephalosporins, yet they have a common reaction mechanisms, which requires the binding of Fe2+, 2-oxoglutarate, O2 and ascorbate at the catalytic site. The two regions of homology, and specifically the identical histidines, potentially represent functionally important sites related to their catalytic activity. Modification of histidine residues by diethyl pyrocarbonate inactivated vertebrate and algal prolyl 4-hydroxylase and vertebrate lysyl hydroxylase, indicating that histidine residues function in the catalytic site of these 2-oxoglutarate-coupled dioxygenases. Inactivation was prevented by the presence of co-substrates, but not by the peptide substrate. It is proposed that the histidine residues in the conserved motifs may function as Fe(2+)-binding ligands.