Development of proteomics on biomodulators as food seed proteins

本论文研究了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两种调节剂。首先，我们分离并鉴定了大豆中编码天冬氨酸蛋白酶（Aps）的两个基因。这些编码的酶，soyAP 1和soyAP 2，共享57%的氨基酸序列同一性。北方印迹分析表明，soyAP 1在种子中特异表达，特别是在干种子中; soyAP 2在根、茎、叶和花等多种组织中表达，但在干种子中不表达。即使在萌发的早期阶段，EAP 1也高度表达，随后表达强度降低。然而，在室温下吸胀48小时后，GSTAP 2 mRNA水平增加。在根尖中，表达于未成熟的导管分子中，当这些分子还未成熟时，这些分子就消失了。本研究从小麦种子中克隆了4种半胱氨酸蛋白酶抑制剂的cDNA，分别为WC 1、WC 2、WC 3和WC 4。所有这些都在成熟的早期阶段在种子中表达，之后，它们的数量逐渐减少。然而，每种mRNA在萌发开始后一天再次表达，并且表达持续接下来的五天。与WC 2和WC 4相比，WC 1似乎总是以更高的水平表达。免疫染色结果表明，在糊粉层和胚中均存在WC 1和WC 4，而在胚乳中仅存在WC 1。在mRNA水平和组织定位的差异发现的WC可能表明其不同的生理作用。

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.