INHIBITION OF PROLYL HYDROXYLATION DURING COLLAGEN BIOSYNTHESIS IN HUMAN-SKIN FIBROBLAST-CULTURES BY ETHYL 3,4-DIHYDROXYBENZOATE
INHIBITION OF PROLYL HYDROXYLATION DURING COLLAGEN BIOSYNTHESIS IN HUMAN-SKIN FIBROBLAST-CULTURES BY ETHYL 3,4-DIHYDROXYBENZOATE
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DOI：
10.1111/1523-1747.ep12471775
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发表时间：
1987-10-01
影响因子：
6.5
通讯作者：
UITTO, J
中科院分区：
文献类型：
--
作者：
MAJAMAA, K;SASAKI, T;UITTO, J
4-羟脯氨酸的酶催化形成在胶原蛋白的细胞内生物合成中起关键作用，因为在生理条件下三螺旋前胶原分子的合成和分泌需要临界数量的4-羟脯氨酸残基。催化脯氨酰残基转化为4-羟脯氨酸的酶，脯氨酰4-羟化酶，需要亚铁离子，α-酮戊二酸盐和抗坏血酸盐。已知3，4-二羟基苯甲酸作为纯化脯氨酰4-羟化酶的有效竞争性抑制剂，与该酶的一种或几种辅因子或辅底物有关。然而，3，4-二羟基苯甲酸是完整细胞中脯氨酰羟基化的不良抑制剂，可能是由于其极性不允许其进入细胞。在这项研究中，在人皮肤成纤维细胞培养物中测试了3，4-二羟基苯甲酸的几种疏水修饰，以确定其抑制4-羟脯氨酸合成的功效。结果表明，3，4-二羟基苯甲酸的乙酯是成纤维细胞培养物中脯氨酰羟基化的有效抑制剂，Ki为约0.4 3，4-二羟基苯甲酸乙酯对赖氨酰残基的羟基化作用很小，不影响细胞总蛋白质合成或DNA复制。为了检验3，4-二羟基苯甲酸乙酯可能用作潜在的抗纤维化剂的假设，还测试了其在硬皮病成纤维细胞中抑制脯氨酰羟基化的功效。结果表明，3，4-二羟基苯甲酸乙酯同样降低了硬皮病细胞培养物中4-羟脯氨酸的合成。因此，脯氨酰4-羟化酶的辅因子或辅底物的结构类似物，如本文测试的3，4-二羟基苯甲酸乙酯或其进一步修饰，也可以在体内用作脯氨酰残基的翻译后羟基化的抑制剂。这些化合物可以潜在地提供减少纤维化疾病（例如硬皮病）中组织中胶原沉积的新方法。
The enzymatically catalyzed formation of 4-hydroxyproline plays a key role in the intracellular biosynthesis of collagen, since a critical number of 4-hydroxyprolyl residues is required for synthesis and secretion of triple-helical procollagen molecules under physiologic conditions. The enzyme catalyzing the conversion of prolyl residues to 4-hydroxyproline, prolyl 4-hydroxylase, requires ferrous ion, .alpha.-ketoglutarate, and ascorbate for its activity. 3,4-Dihydroxybenzoic acid has been known to act as potent competitive inhibitor of purified prolyl 4-hydroxylase with respect to one or several of the cofactors or cosubstrates of the enzyme. 3,4-Dihydroxybenzoic acid, however, is a poor inhibitor of prolyl hydroxylation in intact cells, probably due to its polarity not allowing it to enter the cells. In this study, several hydrophobic modifications of 3,4-dihydroxybenzoic acid were tested in human skin fibroblast cultures for their efficacy to inhibit the synthesis of 4-hydroxyproline. The results indicated that the ethylester of 3,4-dihydroxybenzoic acid was an efficient inhibitor of prolyl hydroxylation in fibroblast cultures, with Ki of .apprx. 0.4 mM. Ethyl 3,4-dihydroxybenzoate had little, if any, effect on the hydroxylation of lysyl residues, andit did not affect total protein synthesis or DNA replication in these cells. To test the hypothesis that ethyl 3,4-dihydroxybenzoate might serve as a potential antifibrotic agent, its efficacy in inhibiting prolyl hydroxylation in scleroderma fibroblasts was also tested. The results indicated that the synthesis of 4-hydroxyproline in scleroderma cell cultures was similarly reduced by ethyl 3,4-dihydroxybenzoate. Thus, structural analogs of the cofactors or cosubstrates of prolyl 4-hydroxylase, such as ethyl 3,4-dihydroxybenzoate tested here or its further modifications, may serve as inhibitors of posttranslational hydroxylation of prolyl residues also in vivo. These compounds could potentially provide a novel means of reducing collagen deposition in tissues in fibrotic diseases, such as scleroderma.