The assessment of the importence of the nearby sequence before peroxisomal targeting sequence (PTS1) on the transport of peroxisomal matrix protein SPS19 in Saccharomyces cerevisiae

• Main Authors: Yu-Chen Yen, 嚴玉真
• Other Authors: 蔡榮宗
• Format: Others
• Language: zh-TW
• Published: 2013
• Online Access: http://ndltd.ncl.edu.tw/handle/49810494097136559460

碩士 === 中山醫學大學 === 生化暨生物科技研究所 === 101 === Peroxisomal matrix proteins are imported from cytoplasm into peroxisomes through peroxisomal targeting signal 1 (PTS1). Peroxisomal targeting signal (PTS) type 1 (PTS1) is a C-terminal uncleaved tripeptide (serine-lysine-leucine [SKL] or variants). Analysis of sequence variability in the PTS1 motif revealed that, in addition to the known C-terminal tripeptide, at least nine residues directly upstream are important for signal recognition in the PTS1:Pex5p receptor complex. The yeast SPS19 gene encoding the peroxisomally targeted 2,4-dienoyl-CoA reductase ，fatty acids with double bonds at even-numbered positions require for their degradation the auxiliary enzyme 2,4-dienoyl-CoA reductase。The Saccharomyces cerevisiae SPS19 deleted was reduction in sporulation and sensitive to lytic.SPS19 reveled a carboxyl-terminal SKL peroxisome targeting signal. In this study , we constructed pRS424-Gal-GFP-PTS1-12a，pRS424-Gal-GFP-PTS1-3ａ, and pRS424-Gal-GFP, transformed these plasmid into wild-type W303.1a and W303.1a-Δpex19 by yeast transformation technique .The GFP-PTS1 encoding these strains were assayed peroxisome formation by the fluorescence microscopy technique.pRS424-Gal-GFP-PTS1-12a and pRS424-Gal-GFP-PTS1-3a DNA sequence use ClustalW2 analysis to perform comparative sequencing .We find the expected results for the pRS424-Gal-GFP-PTS1-20a and pRS424-Gal-GFP-PTS1-12a have green fluorescence in peroxisome, but the pRS424-Gal-GFP-PTS1-3a has green fluorescence less than the pRS424-Gal-GFP-PTS1-20a and pRS424-Gal-GFP-PTS1-12a in peroxisome, and pRS424-Gal-GFP have green fluorescence in cytosol. These results suggest that the C-terminal 12 amino acids of SPS19 is better sequence reguired for effective protein transport into peroxisome.