Cloning and chromosomal mapping of the human nonfunctional gene for L-gulono-gamma-lactone oxidase, the enzyme for L-ascorbic acid biosynthesis missing in man.
Cloning and chromosomal mapping of the human nonfunctional gene for L-gulono-gamma-lactone oxidase, the enzyme for L-ascorbic acid biosynthesis missing in man.
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DOI：
10.1016/s0021-9258(17)36884-9
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发表时间：
1994-05
期刊：
影响因子：
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通讯作者：
M. Nishikimi;R. Fukuyama;S. Minoshima;N. Shimizu;K. Yagi
中科院分区：
文献类型：
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作者：
M. Nishikimi;R. Fukuyama;S. Minoshima;N. Shimizu;K. Yagi
人类是无法合成l -抗坏血酸的特殊高等动物之一，因为他们缺乏l -古罗酮- γ -内酯氧化酶，这种酶催化l -抗坏血酸生物合成的最后一步。在本研究中，我们从人类基因组文库中分离出一段无功能的l -古罗诺- γ -内酯氧化酶基因，并利用人类染色体流分选和荧光原位杂交技术进行斑点杂交，将其定位在染色体8p21.1上。测序分析表明，分离的片段代表基因的3'部分，其中大鼠l -古洛诺- γ -内酯氧化酶基因的外显子VII、IX、X和XII对应的区域保留，而外显子VIII和XI对应的区域可能缺失。在所鉴定的外显子区域发现了各种异常的核苷酸变化，如核苷酸的缺失和插入，以及在内含子/外显子边界不符合GT/AG规则。将这4个外显子序列推导出的概念氨基酸序列与大鼠的相应序列进行比较，发现存在大量的非保守性替换和2个终止密码子。这些发现表明，人类无功能的l -古罗诺- γ -内酯氧化酶基因在进化过程中停止了功能，在没有选择压力的情况下积累了大量突变。
Man is among the exceptional higher animals that are unable to synthesize L-ascorbic acid because of their deficiency in L-gulono-gamma-lactone oxidase, the enzyme catalyzing the terminal step in L-ascorbic acid biosynthesis. In the present study, we isolated a segment of the nonfunctional L-gulono-gamma-lactone oxidase gene from a human genomic library, and mapped it on chromosome 8p21.1 by spot blot hybridization using flow-sorted human chromosomes and fluorescence in situ hybridization. Sequencing analysis indicated that the isolated segment represented a 3'-part of the gene, where the regions corresponding to exons VII, IX, X, and XII of the rat L-gulono-gamma-lactone oxidase gene remain with probable deletion of the regions corresponding to exons VIII and XI. In the identified exon regions were found various anomalous nucleotide changes, such as deletion and insertion of nucleotide(s) and nonconformance to the GT/AG rule at intron/exon boundaries. When the conceptual amino acid sequences deduced from the four exon sequences were compared with the corresponding rat sequences, there were a large number of nonconservative substitutions and also two stop codons. These findings indicate that the human nonfunctional L-gulono-gamma-lactone oxidase gene has accumulated a large number of mutations without selective pressure since it ceased to function during evolution.