Shan X, Xue Z, Mélèse T
Department of Biological Sciences, Columbia University, New York 10027.
J Cell Biol. 1994 Aug;126(4):853-62. doi: 10.1083/jcb.126.4.853.
We have identified a gene (NPI46) encoding a new prolyl cis-trans isomerase within the nucleolus of the yeast Saccharomyces cerevisiae. The protein encoded by NPI46 was originally found by us in a search for proteins that recognize nuclear localization sequences (NLSs) in vitro. Thus, NPI46 binds to affinity columns that contain a wild-type histone H2B NLS but not a mutant H2B NLS that is incompetent for nuclear localization in vivo. NPI46 has two domains, a highly charged NH2 terminus similar to two other mammalian nucleolar proteins, nucleolin and Nopp140, and a COOH terminus with 45% homology to a family of mammalian and yeast proline isomerases. NPI46 is capable of catalyzing the prolyl cis-trans isomerization of two small synthetic peptides, succinyl-Ala-Leu-Pro-Phe-p-nitroanilide and succinyl-Ala-Ala-Pro-Phe-p-nitroanilide, as measured by a chymotrypsin-coupled spectrophotometric assay. By indirect immunofluorescence we have shown that NPI46 is a nucleolar protein. NPI46 is not essential for cell viability.
我们在酿酒酵母的核仁中鉴定出了一个编码新型脯氨酰顺反异构酶的基因(NPI46)。我们最初是在寻找体外识别核定位序列(NLSs)的蛋白质时发现了由NPI46编码的蛋白质。因此,NPI46能与含有野生型组蛋白H2B NLS的亲和柱结合,但不能与体内无核定位能力的突变型H2B NLS结合。NPI46有两个结构域,一个高度带电的NH2末端,类似于另外两种哺乳动物核仁蛋白——核仁素和Nopp140,还有一个COOH末端,与哺乳动物和酵母脯氨酸异构酶家族有45%的同源性。通过胰凝乳蛋白酶偶联分光光度法测定,NPI46能够催化两种小的合成肽——琥珀酰 - 丙氨酸 - 亮氨酸 - 脯氨酸 - 对硝基苯胺和琥珀酰 - 丙氨酸 - 丙氨酸 - 脯氨酸 - 对硝基苯胺的脯氨酰顺反异构化。通过间接免疫荧光法,我们证明NPI46是一种核仁蛋白。NPI46对细胞活力并非必需。