油菜花粉钙结合蛋白的研究

Studies on Calcium Binding Proteins in Brassica Campestris Pollen

以油菜花粉（Ｂｒａｓｉｃａｃａｍｐｅｓｔｒｉｓ）为材料纯化了钙调素（ＣａＭ），并得到一种新钙结合蛋白（ＮＣＢＰ）。经ＳＤＳＰＡＧＥ、ＰＡＧＥ和等电聚焦电泳（ＩＥＦ）鉴定，这两种蛋白质均表现均一。ＣａＭ分子量为１８８ｋＤ，ＮＣＢＰ为１６３ｋＤ，等电点分别为３６和４２。研究证明花粉ＣａＭ具有与其他来源ＣａＭ所特有的性质，对环核苷酸磷酸二酯酶（简称ＰＤＥ）有明显的激活作用，而花粉ＮＣＢＰ不明显。花粉ＣａＭ在ＰＡＧＥ和ＳＤＳＰＡＧＥ中电泳行为有Ｃａ２＋效应，而ＮＣＢＰ仅表现在ＰＡＧＥ中。经氨基酸组成分析表明两种蛋白质氨基酸组成不同，但均含有较多的酸性氨基酸，不含Ｔｒｐ，含１个Ｃｙｓ。ＣａＭ和ＮＣＢＰＮ端均为封闭，ＣａＭＣ端为ＭｅｔＡｌａＬｙｓＣＯＯＨ。两种蛋白质具有不同的肽谱和ＣＤ谱。用ＤＴＮＢ修饰花粉ＣａＭＣｙｓ残基，则激活ＰＤＥ的能力明显下降。

Both Calmodulin(CaM) and a Neo Calcium Binding Protein(NCBP) were purified from Brassica campestris pollen by phenyl Sepharose CL 4B hydrophobic chromato graphy and DEAE F.F.ion exchange chromatography.Two proteins were shown to be h omogeneous by SDS PAGE, PAGE and IEF.Molecular weights and pI points of pollen CaM and NCBP are 18.8 and 16.3 kD,3.6 and 4.2 respectively. Pollen CaM can activ ate bovine heart cyclic nucleotide phosphodiesterase, pollen NCBP is not notable . Pollen CaM shows Calcium ion effect in both SDS PAGE and PAGE, while NCBP onl y does in PAGE. The amino acid composition of pollen CaM and NCBP is different, b o th contain more acidic amino acids and a Cys, but lack Trp.Both of their N term inus are blocked and C terminus of pollen CaM is Met Ala Lys COOH.Their pe p tide maps in HPLC and CD spectrums are different. The Cys of pollen CaM was modi f ied by DTNB and the PDE activity by CaM activation is marked descent. From above we can conclude that pollen NCBP is a new Calcium Binding Protein different from pollen CaM.