Höhfeld J, Veenhuis M, Kunau W H
Abteilung für Zellbiochemie, Ruhr-Universität Bochum, Federal Republic of Germany.
J Cell Biol. 1991 Sep;114(6):1167-78. doi: 10.1083/jcb.114.6.1167.
Saccharomyces cerevisiae pas3-mutants are described which conform the pas-phenotype recently reported for the peroxisomal assembly mutants pas1-1 and pas2 (Erdmann, R., M. Veenhuis, D. Mertens, and W.-H Kunau, 1989, Proc. Natl. Acad. Sci. USA. 86:5419-5423). The isolation of pas3-mutants enabled us to clone the PAS3 gene by functional complementation. DNA sequence analysis revealed a 50.6-kD protein with at least one domain of sufficient length and hydrophobicity to span a lipid bilayer. To verify these predictions antibodies were raised against a truncated portion of the PAS3 coding region overexpressed in E. coli. Pas3p was identified as a 48 kD peroxisomal integral membrane protein. It is shown that a lack of this protein causes the peroxisome-deficient phenotype and the cytosolic mislocalization of peroxisomal matrix enzymes. Based on protease digestion experiments Pas3p is discussed to be anchored in the peroxisomal membrane by its amino-terminus while the bulk of the molecule is exposed to the cytosol. These findings are consistent with the possibility that Pas3p is one component of the peroxisomal import machinery.
已描述了酿酒酵母pas3突变体，其符合最近报道的过氧化物酶体组装突变体pas1-1和pas2的pas表型（埃德曼，R.，M. 费恩胡伊斯，D. 默滕斯，以及W.-H. 库瑙，1989，《美国国家科学院院刊》。86:5419 - 5423）。pas3突变体的分离使我们能够通过功能互补克隆PAS3基因。DNA序列分析揭示了一种50.6-kD的蛋白质，其具有至少一个足够长且具疏水性的结构域以跨越脂质双层。为验证这些预测，针对在大肠杆菌中过表达的PAS3编码区的截短部分制备了抗体。Pas3p被鉴定为一种48 kD的过氧化物酶体整合膜蛋白。结果表明，这种蛋白质的缺失会导致过氧化物酶体缺陷表型以及过氧化物酶体基质酶的胞质错误定位。基于蛋白酶消化实验，讨论了Pas3p通过其氨基末端锚定在过氧化物酶体膜中，而分子的大部分暴露于胞质溶胶中。这些发现与Pas3p是过氧化物酶体输入机制的一个组成部分的可能性一致。
