Wu M, Wong S M, Tan H M, Ting R
Institute of Molecular and Cell Biology, National University of Singapore.
Biochem Biophys Res Commun. 1995 Oct 13;215(2):578-90. doi: 10.1006/bbrc.1995.2504.
A respiratory defective pet mutant (W303 delta FUM1) of Saccharomyces cerevisiae deficient in fumarase was transformed with the plasmid construct pG5/ST7. This plasmid contains the entire FUM1 gene with only a single base pair change (ATG-->ATC) confined to the putative second inframe translation initiation codon. While transformation of the fumarase deficient mutant with pG5/ST7 resulted in an elevation of fumarase activity in the mitochondria of the transformed strain, fumarase activity in the cytosol remained negligible his result indicated that the cytosolic fumarase isoenzyme is synthesized exclusively from the second translation initiation codon of FUM1. Results of RACE-PCR of the 5' ends of FUM1 transcripts confirmed that the FUM1 gene synthesizes two distinct species of transcripts. These data provide strong evidence for the synthesis and targeting of two fumarase isomers.
用质粒构建体pG5/ST7转化了酿酒酵母中缺乏延胡索酸酶的呼吸缺陷型宠物突变体（W303 delta FUM1）。该质粒包含完整的FUM1基因，只有一个碱基对变化（ATG-->ATC）限于推定的第二个框内翻译起始密码子。当用pG5/ST7转化延胡索酸酶缺陷型突变体时，转化菌株线粒体中的延胡索酸酶活性升高，而细胞质中的延胡索酸酶活性仍然可以忽略不计。这一结果表明，细胞质延胡索酸酶同工酶仅从FUM1的第二个翻译起始密码子合成。FUM1转录本5'端的RACE-PCR结果证实，FUM1基因合成两种不同的转录本。这些数据为两种延胡索酸酶异构体的合成和靶向提供了有力证据。
