Structural characterization of the gene and corresponding cDNA for the cytochrome P450rm from Rhodotorula minuta which catalyzes formation of isobutene and 4-hydroxylation of benzoate
Structural characterization of the gene and corresponding cDNA for the cytochrome P450rm from Rhodotorula minuta which catalyzes formation of isobutene and 4-hydroxylation of benzoate
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DOI：
10.1007/s004380050552
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发表时间：
1997-09-01
期刊：
影响因子：
--
通讯作者：
Fukuda, H
中科院分区：
文献类型：
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作者：
Fujii, T;Nakamura, K;Fukuda, H
细胞色素P450rm是一种双功能酶，具有异丁烯形成酶和苯甲酸酯4-羟化酶活性。我们克隆了P450rm的基因和相应的cDNA，以便在真菌系统发育和生理学的背景下表征该酶。从cDNA序列推断，P450rm有527个氨基酸，计算分子量为59 136。P450rm与黑曲霉CYP53A1的氨基酸序列同源性为48%，表明该基因属于CYP53的一个新亚家族CYP53B。然而，P450rm基因具有8个外显子和7个内含子，其结构与CYP53A1完全不同。Northern分析表明，当l -苯丙氨酸作为唯一碳源时，P450rm mRNA的表达水平升高。这些结果表明，在真菌从l -苯丙氨酸开始的异化途径中，P450rm作为苯甲酸酯4-羟化酶在进化过程中得到了很好的保存。
Cytochrome P450rm was previously isolated from the basidiomycete yeast Rhodotorula minuta as a bifunctional enzyme with isobutene-forming and benzoate 4-hydroxylase activities. We cloned the gene and corresponding cDNA for P450rm in order to characterize the enzyme in the context of fungal phylogeny and physiology. From the cDNA sequence, P450rm was deduced to have 527 amino acids with a calculated molecular weight of 59 136. P450rm shared 48% amino acid sequence identity with CYP53A1 from Aspergillus niger, indicating that the gene belongs to a novel subfamily of CYP53, CYP53B. However, the organization of the P450rm gene, which has eight exons and seven introns, differed completely to that of CYP53A1. Northern analysis demonstrated that the level of P450rm mRNA expression increased when L-phenylalanine was used as sole carbon source. These results suggest that P450rm has been well conserved during the evolution of fungi as a benzoate 4-hydroxylase in the dissimilation pathway starting from L-phenylalanine.