Kihara Akihiko, Saburi Wataru, Wakuta Shinji, Kim Myung-Hee, Hamada Shigeki, Ito Hiroyuki, Imai Ryozo, Matsui Hirokazu
Laboratory of Biochemistry, Research Faculty of Agriculture, Hokkaido University.
Biosci Biotechnol Biochem. 2011;75(9):1740-5. doi: 10.1271/bbb.110285. Epub 2011 Sep 7.
Nucleoside diphosphate kinase (NDPK) is a ubiquitous enzyme that catalyzes the transfer of the γ-phosphoryl group from a nucleoside triphosphate to a nucleoside diphosphate. In this study, we examined the subcellular localization, tissue-specific gene expression, and enzymatic characteristics of three rice NDPK isozymes (OsNDPK1-OsNDPK3). Sequence comparison of the three OsNDPKs suggested differential subcellular localization. Transient expression of green fluorescence protein-fused proteins in onion cells indicated that OsNDPK2 and OsNDPK3 are localized to plastid and mitochondria respectively, while OsNDPK1 is localized to the cytosol. Expression analysis indicated that all the OsNDPKs are expressed in the leaf, leaf sheath, and immature seeds, except for OsNDPK1, in the leaf sheath. Recombinant OsNDPK2 and OsNDPK3 showed lower optimum pH and higher stability under acidic pH than OsNDPK1. In ATP formation, all the OsNDPKs displayed lower K(m) values for the second substrate, ADP, than for the first substrate, NTP, and showed lowest and highest K(m) values for GTP and CTP respectively.
核苷二磷酸激酶（NDPK）是一种普遍存在的酶，它催化γ-磷酸基团从核苷三磷酸转移至核苷二磷酸。在本研究中，我们检测了三种水稻NDPK同工酶（OsNDPK1 - OsNDPK3）的亚细胞定位、组织特异性基因表达及酶学特性。三种OsNDPK的序列比较表明其亚细胞定位存在差异。绿色荧光蛋白融合蛋白在洋葱细胞中的瞬时表达表明，OsNDPK2和OsNDPK3分别定位于质体和线粒体，而OsNDPK1定位于细胞质溶胶。表达分析表明，除OsNDPK1在叶鞘中不表达外，所有OsNDPK均在叶片、叶鞘和未成熟种子中表达。重组OsNDPK2和OsNDPK3在酸性pH条件下显示出比OsNDPK1更低的最适pH和更高的稳定性。在ATP形成过程中，所有OsNDPK对第二底物ADP的K(m)值均低于对第一底物NTP的K(m)值，并且对GTP和CTP分别显示出最低和最高的K(m)值。
