Metzner M, Stoller G, Rücknagel K P, Lu K P, Fischer G, Luckner M, Küllertz G
Institut für Pharmazeutische Biologie, Martin-Luther-Universität Halle-Wittenberg, Hoher Weg 8, D-06120 Halle/Saale, Germany.
J Biol Chem. 2001 Apr 27;276(17):13524-9. doi: 10.1074/jbc.M007005200. Epub 2000 Dec 15.
A functionally Pin1-like peptidyl-prolyl cis/trans isomerase (PPIase(1)) was isolated from proembryogenic masses (PEMs) of Digitalis lanata according to its enzymatic activity. Partial sequence analysis of the purified enzyme (DlPar13) revealed sequence homology to members of the parvulin family of PPIases. Similar to human Pin1 and yeast Ess1, it exhibits catalytic activity toward substrates containing (Thr(P)/Ser(P))-Pro peptide bonds and comparable inhibition kinetics with juglone. Unlike Pin1-type enzymes it lacks the phosphoserine or phosphothreonine binding WW domain. Western blotting with anti-DlPar13 serum recognized the endogenous form in nucleic and cytosolic fractions of the plant cells. Since the PIN1 homologue ESS1 is an essential gene, complementation experiments in yeast were performed. When overexpressed in Saccharomyces cerevisiae DlPar13 is almost as effective as hPin1 in rescuing the temperature-sensitive phenotype caused by a mutation in ESS1. In contrast, the human parvulin hPar14 is not able to rescue the lethal phenotype of this yeast strain at nonpermissive temperatures. These results suggest a function for DlPar13 rather similar to parvulins of the Pin1-type.
根据其酶活性，从毛花洋地黄的胚性细胞团（PEMs）中分离出一种功能上类似于Pin1的肽基脯氨酰顺/反异构酶（PPIase(1)）。对纯化酶（DlPar13）的部分序列分析显示，其与PPIase小菌素家族成员具有序列同源性。与人类Pin1和酵母Ess1相似，它对含有（苏氨酸磷酸化/丝氨酸磷酸化）-脯氨酸肽键的底物表现出催化活性，并且与胡桃醌具有相似的抑制动力学。与Pin1型酶不同，它缺乏磷酸丝氨酸或磷酸苏氨酸结合的WW结构域。用抗DlPar13血清进行的蛋白质印迹法在植物细胞的细胞核和细胞质组分中识别出内源性形式。由于PIN1同源物ESS1是一个必需基因，因此在酵母中进行了互补实验。当在酿酒酵母中过表达时，DlPar13在挽救由ESS1突变引起的温度敏感表型方面几乎与hPin1一样有效。相比之下，人类小菌素hPar14在非允许温度下无法挽救该酵母菌株的致死表型。这些结果表明DlPar13的功能与Pin1型小菌素相当相似。
