Abel S, Nürnberger T, Ahnert V, Krauss G J, Glund K
Department of Vegetable Crops, University of California, Davis, California 95616, USA.
Plant Physiol. 2000 Feb;122(2):543-52. doi: 10.1104/pp.122.2.543.
During growth under conditions of phosphate limitation, suspension-cultured cells of tomato (Lycopersicon esculentum Mill.) secrete phosphodiesterase activity in a similar fashion to phosphate starvation-inducible ribonuclease (RNase LE), a cyclizing endoribonuclease that generates 2':3'-cyclic nucleoside monophosphates (NMP) as its major monomeric products (T. Nürnberger, S. Abel, W. Jost, K. Glund [1990] Plant Physiol 92: 970-976). Tomato extracellular phosphodiesterase was purified to homogeneity from the spent culture medium of phosphate-starved cells and was characterized as a cyclic nucleotide phosphodiesterase. The purified enzyme has a molecular mass of 70 kD, a pH optimum of 6.2, and an isoelectric point of 8.1. The phosphodiesterase preparation is free of any detectable deoxyribonuclease, ribonuclease, and nucleotidase activity. Tomato extracellular phosphodiesterase is insensitive to EDTA and hydrolyzes with no apparent base specificity 2':3'-cyclic NMP to 3'-NMP and the 3':5'-cyclic isomers to a mixture of 3'-NMP and 5'-NMP. Specific activities of the enzyme are 2-fold higher for 2':3'-cyclic NMP than for 3':5'-cyclic isomers. Analysis of monomeric products of sequential RNA hydrolysis with purified RNase LE, purified extracellular phosphodiesterase, and cleared -Pi culture medium as a source of 3'-nucleotidase activity indicates that cyclic nucleotide phosphodiesterase functions as an accessory ribonucleolytic activity that effectively hydrolyzes primary products of RNase LE to substrates for phosphate-starvation-inducible phosphomonoesterases. Biosynthetical labeling of cyclic nucleotide phopshodiesterase upon phosphate starvation suggests de novo synthesis and secretion of a set of nucleolytic enzymes for scavenging phosphate from extracellular RNA substrates.
在磷限制条件下生长时，番茄（Lycopersicon esculentum Mill.）悬浮培养细胞分泌磷酸二酯酶活性的方式与磷饥饿诱导的核糖核酸酶（RNase LE）相似，RNase LE是一种环化内切核糖核酸酶，其主要单体产物是2':3'-环核苷单磷酸（NMP）（T. Nürnberger、S. Abel、W. Jost、K. Glund [1990] Plant Physiol 92: 970 - 976）。从磷饥饿细胞的废培养基中纯化出番茄细胞外磷酸二酯酶至同质，并将其鉴定为环核苷酸磷酸二酯酶。纯化后的酶分子量为70 kD，最适pH为6.2，等电点为8.1。该磷酸二酯酶制剂不含任何可检测到的脱氧核糖核酸酶、核糖核酸酶和核苷酸酶活性。番茄细胞外磷酸二酯酶对EDTA不敏感，且无明显碱基特异性地将2':3'-环NMP水解为3'-NMP，将3':5'-环异构体水解为3'-NMP和5'-NMP的混合物。该酶对2':3'-环NMP的比活性比对3':5'-环异构体高2倍。用纯化的RNase LE、纯化的细胞外磷酸二酯酶以及作为3'-核苷酸酶活性来源的澄清-Pi培养基对RNA连续水解的单体产物进行分析表明，环核苷酸磷酸二酯酶作为一种辅助核糖核酸水解活性，可有效地将RNase LE的初级产物水解为磷饥饿诱导的磷酸单酯酶的底物。磷饥饿时环核苷酸磷酸二酯酶的生物合成标记表明，为从细胞外RNA底物中清除磷，会从头合成并分泌一组核酸水解酶。
