Development of proteomics on biomodulators as food seed proteins

对大豆中的天冬氨酸蛋白酶和水稻种子中的半胱氨酸蛋白酶抑制剂进行了研究。首先，我们从大豆甘氨酸中分离并鉴定了两个编码天冬氨酸蛋白酶（Aps）的基因。所编码的酶soyAP1和soyAP2具有57%的氨基酸序列同源性。Northern blot分析表明，soyAP1在种子中特异性表达，特别是在干种子中，而soyAP2在根、茎、叶、花等多种组织中均有表达，但在干种子中不表达。大豆ap1在萌发初期也有高表达，随后表达强度下降。室温渗吸48 h后，SoyAP2 mRNA水平升高。在根尖处，SoyAP1在未成熟气管元件中表达，在元件未成熟时消失。这些结果表明大豆ap1在未成熟气管元件和筛管中表达，并可能参与细胞程序性死亡。我们从小麦种子中克隆了4种小麦胱抑素的cdna，分别是WC1、WC2、WC3和WC4。这些基因均在种子成熟初期表达，成熟后表达量逐渐减少。然而，每个mrna在发芽开始后一天再次表达，并在接下来的五天内持续表达。与WC2和WC4相比，WC1的表达水平似乎始终较高。免疫染色研究表明，WC1和WC4均存在于糊层和胚中，而在胚乳中仅存在WC1。WCs的mRNA水平和组织定位的差异可能表明它们的不同生理作用。

We have studied about two modulations : aspartic proteinase in soybean and cysteine proteinase inhibitor in rice seeds. First, we isolated and characterized two genes coding for aspartic proteinases (Aps) in soybean Glycine max. These encoded enzymes, soyAP1 and soyAP2, shared 57% amino acid sequence identity. Northern blot analysis demonstrated that soyAP1 was expressed specifically in seeds, especially in dry seeds, while the expression of soyAP2 took place in various tissues such as roots, stems, leaves and flowers, but not in dry seeds. SoyAP1 was highly expressed even at an early stage of germination, with a subsequent decrease in expression intensity. However, SoyAP2 mRNA level increased when 48 hours elapsed after imbibition at room temperature. At the root tip, SoyAP1 was expressed in immature tracheary elements disappeared when the elements were still immature. These results suggest that soyAP1 is expressed in immature tracheary elements and sieve tubes, and may be involved in programmed cell death.We cloned four kinds of cDNAs for wheat cystatins, WC1, WC2, WC3 and WC4, from the seed. All were expressed in the seed at an early stage of maturation and, after that, their quantities decreased gradually. However, each of the mRNAs was again expressed one day after the start of germination and the expression continued for the following five days. WC1 seemed to be expressed always at higher level compared to WC2 and WC4. Immunostaining study showed that both WC1 and WC4 existed in the aleuron layer and embryo, whereas in the endosperm the only existing species was WC1. Differences in mRNA level and tissue localization found for the WCs may suggest their differential physiological roles.