La3+ Induced Binding of Calmodulin(CaM) to CaM-binding Proteins in Rat Brain Homogenate

应用固定化钙调蛋白(CaM)亲合色谱法、变性丙烯酰胺电泳(SDS-PAGE)和基质辅助激光解析电离飞行时间质谱(MALDI-TOF-MS)等方法研究了La3+诱导CaM在大鼠脑匀浆液中的钙调蛋白亲合蛋白(CaMBP)谱以及CaM-CaMBP在模拟细胞环境下结合作用的La3+浓度依赖关系,并与Ca2+的作用进行了比较.实验结果表明,(1)La3+参与的CaMBP物种与Ca2+的基本相同.鉴定了其中含量高且稳定出现的5种CaMBP分别是参与糖酵解反应的6-磷酸果糖激酶和3-磷酸甘油醛脱氢酶、细胞骨架类的微管蛋白和肌动蛋白以及应激反应相关的71000热休克同源蛋白,表明稀土离子可能参与多种细胞过程;...Interactions of lanthanide ions with Ca2+/calmodulin (CaM) signal transduction is a crucial pathway way for the biological effect of lanthanides. In the present work, the effect of La3+ on binding of CaM to its binding proteins (CaMBP) was investigated by an immobilized CaM affinity chromatographic method in rat's brain homogenate. CaMBP binding to La3+-activated CaM gel were analyzed by SDS-PAGE and some CaMBP were identified by MALDI-TOF-MS. The concentration dependency on La3+ for CaM-CaMBP binding was determined in a mimetic cellular media and compared with that of Ca2+. The experimental results showed that (1) the CaMBP species activated by La3+ were primarily the same as those of Ca2+. Five selected CaMBP were identified to be 6-phosphofructokinase (PFK), glyceraldehyde-3-phosphate dehydrogenase (GAPDH) (enzymes in glycolysis), tubulin, actin (cytoskeleton proteins), and heat shock cognate 71000 protein (HSCP71, stress chaperone), indicating that La3+ may interfere with multiple cellular processes through Cam. (2) The effective concentrations for La3+ to induce binding of CaM to CaMBP depended on CaMBP species Tubulin, actin and HSCP71 were observed to be sensitive to La3+ revealed by the affinity constants K of La3+ to CaM in the metal-CaM-CaMBP tertiary system, which are similar or higher than those of Ca2+. However, in the systems with PFK or GAPDH, La3+ bound to CaM less tightly. We hypothesized that this may attribute tribute to complexity of mimetic cellular media and/or the effects of CaM-CaMBP interaction on CaM-metal binding, on which further studies should be appropriate. The work may offer new insight to the mechanisms of the biological effects of lanthanides.http://gateway.webofknowledge.com/gateway/Gateway.cgi?GWVersion=2&SrcApp=PARTNER_APP&SrcAuth=LinksAMR&KeyUT=WOS:000262006900039&DestLinkType=FullRecord&DestApp=ALL_WOS&UsrCustomerID=8e1609b174ce4e31116a60747a720701SCI(E)中文核心期刊要目总览(PKU)中国科技核心期刊(ISTIC)中国科学引文数据库(CSCD)3122525-25302