Hydrolytic Enzyme Inhibitors and Necrotic Reactions in Potato Leaves Reduce Reproductive Success of Colorado Potato Beetle

In field and laboratory conditions the reproductive potential （survival at different stages of development） of the Colorado potato beetle has been measured, the dependence of the parameters of insects fertility on varietal features of the feed has been investigated, the food activity of the representatives of insects from four areas of the Southern Urals has been measured, the intensity and time of hypersensitivity reactions on potato leaves has been defined. In laboratory experiments, the level of activity of hydrolytic enzymes in insects and the level of activity of these enzymes inhibitors in the leaves of three potato varieties differing in their resistance to the Colorado potato beetle have been determined. The results show that the insects-representatives of different samples have different food activity in relation to different potato varieties. High mortality rates were revealed at feeding with leaves of potato of the Bashkir variety characterized by a high level of hydrolases inhibitors and relatively rapid development of necrotic reactions after the appearance of eggs laying on the leaves. We suppose that the neerogenetic barrier appearing in the form of necrosis is the major cause of the fetal mortality of insects. The level of the content of inhibitors inactivating larvae digestive enzymes affects the survival of insects at postembryonic stages. The speed of the necrosis emergence, the level of activity of digestive enzymes inhibitors in potato can be considered as criteria for the selection of breeding material for resistance to the Colorado potato beetle.     

Study on Enzymatic Hydrolysis of Gadus morrhua Skin Collagen and Molecular Weight Distribution of Hydrolysates

Process parameters on enzymatic hydrolysis and molecular weight （MW） distribution of collagen hydrolysates from Gadus morrhua skin were investigated. The optimal process parameters were obtained by the single-factor and orthogonal experiments. The molecular weight distribution of hydrolysates was determined using both Sephadex G25 partition and high speed liquid chromatography electricity spray mass spectrum （HPLC-ESI-MS）. Collagen hydrolysates were first gained by an alkaline protease ＂alcalase＂ for 3 h at temperature （50~C）, pH （10.0）, substrate concentration （75 g L-~）, and E/S （3%）. The molecular weight distribution of collagen hydrolysates ranged from 300 to 1 500 Da, and most of peptides were under 1 200 Da. Sephadex G25 partition and HPLC-ESI-MS should be successfully employed to determine the molecular weight distribution of collagen hydrolysates.     

酶法提取芦笋皮中高活性膳食纤维的研究

以芦笋皮为原料，采用酶法水解制备高活性膳食纤维。试验结果表明：用0.54%的脂肪酶(100 U/mg)、0.80%的淀粉酶(25 U/mg)和0.80%的糖化酶(20 U/mg)、1.20%的蛋白酶(100 U/mg)可提取出高活性的膳食纤维。所得膳食纤维纯度高，总膳食纤维含量达83.08%，其中水溶性膳食纤维17.02%，不溶性膳食纤维66.06%。而且生理活性好，持水性和溶胀性分别为886%和4.5 mL/g。     

种子活力与发芽环境对玉米种子蛋白酶α-淀粉酶活性的影响

为探讨不同活力玉米种子田间发芽成苗率存在明显差异的生理原因,测定了不同环境条件下玉米种子发芽期间蛋白酶和α-淀粉酶的活性.结果表明,种子活力对玉米籽粒蛋白酶和α-淀粉酶活性的影响因发芽环境条件而异.高活力种子胚乳的蛋白酶活性在较低的温度下发芽比低活力种子的高,且在不同温度条件下的变化较小.胚乳蛋白酶活性在10℃和25℃下的比值与种子活力密切相关.玉米种子胚中蛋白酶的活性比胚乳的高,且高活力种子胚的蛋白酶活性较高.α-淀粉酶的活动开始于吸水约5 h时的种子中,发芽5～6 d时活性达最高.在10℃发芽时,高活力种子α-淀粉酶活性显著地高于低活力种子,但在25℃发芽时两者差异不显著.在不良环境条件下发芽,玉米种子的蛋白酶活性增高,α-淀粉酶的活性降低.高活力种子的蛋白酶和α-淀粉酶的稳定性和协调性较好.     

鸭肝蛋白酶的分离纯化及其部分性质研究

经Tris-HCl缓冲液抽提、硫酸铵分级沉淀、DEAE-Sepharose Fast Flow离子交换层析、Sephacryl S-200凝胶过滤层析,得到鸭肝蛋白酶电泳纯制品．纯化倍数为650.16倍,回收率为19.23%．经SDS-PAGE和凝胶过滤测得该酶的分子量为35kD．该酶在pH6.0～11.0,温度低于25℃稳定,当温度上升到55℃时活性迅速下降．该酶在pH10和60℃时达到最高活性．进一步研究得知：Cu^2＋离子对该酶有明显促进作用,而Mn^2＋却使酶几乎失活．以酪蛋白为底物,在pH10.0、37℃时测得米氏常数Km值为1.33%．     

水相酶法提取蛋黄油工艺的响应面分析法研究

采用蛋白酶水解鲜鸡蛋蛋黄提取蛋黄油,通过响应曲面分析法建立蛋黄油提取率的二次多项数学模型,并考察酶添加比例、底物质量浓度和反应时间对蛋黄油提取率的影响,优化出蛋黄油提取率达到56.81%的工艺参数:酶添加比例为2.04%,底物质量浓度为20.28%,反应时间为14 h。     

鸡肉蛋白酶水解工艺条件的研究

为探索鸡肉酶水解工艺条件及其动力学特性，分别采用木瓜蛋白酶、胃蛋白酶、中性蛋白酶、菠萝蛋白酶对鸡肉进行了水解试验，得出最优酶为木瓜蛋白酶；由此选用木瓜蛋白酶做正交试验，考察水解温度、时间、加酶量、pH值和固液比5个因素对酶水解的影响，并确定出最优水解条件为温度50℃，时间7 h，加酶量2.4%(以鸡肉的质量百分数计)，pH 7.0，固液比1∶4；在此条件下，水解度可达26.07%。在此基础上由试验数据推导出描述木瓜蛋白酶水解鸡肉的动力学方程，可为鸡肉酶解生化反应器的设计和鸡肉水解蛋白液或蛋白粉的开发提供一定的理论依据。     

Evidence of peptides in low-molecular-weight fractions of soil organic matter

Summary Organic matter was extracted from three soils, a Berwick sandy loam, a Franklin loamy sand, and a Cumberland silty loam. The extracts were separated into high (>8000 daltons) and low-molecular-weight (<8000 daltons) fractions using gel filtration. Reverse-phase high performance liquid chromatography at 214 nm was used to separate the peptides into low-molecular-weight fractions. Peptide samples were collected with an integrated fraction collector and hydolyzed with an immobilized protease column reactor. High performance liquid chromatography with fluorescence detection was used to determine the amino-acid contents of the collected samples. The results indicated that peptide intermediates are present in soil size fractions. Greater quantities of several amino acids were released from the peptide hydrolyzates of the Berwick sandy loam and Franklin loamy sand, compared with the Cumberland silty loam, an uncultivated soil. These findings indicate that organic intermediates (e.g., peptides) are more prevalent in biologically active soils than in relatively inert soils.     

月季花瓣丝氨酸蛋白酶基因RhSep1的克隆及生物信息学分析

利用RT-PCR技术,从切花月季品种"Samantha"中克隆丝氨酸蛋白酶基因RhSep1。利用生物信息学技术对所得到的丝氨酸蛋白酶RhSep1序列进行结构与功能预测。结果表明:RhSep1基因全长2 442 bp,开放阅读框编码769个氨基酸,推定该丝氨酸蛋白酶分子质量为80.48 kD。通过NCBI和MEROPS肽酶数据库等对Rh-Sep1进行Protein Blast,发现RhSep1具有肽酶S8₃家族SA的典型结构域Petidase_S8₃（序列为Tyr-108-Leu-341）和PA_subtilisin_like的结构域（序列为Tyr-348-Ile-474）。预测RhSep1可能具有信号肽、明显疏水区和典型跨膜区,同时RhSep1氨基酸序列中存在较多蛋白激酶C和酪蛋白激酶Ⅱ磷酸化位点、豆蔻酰化位点。这些活性位点往往与蛋白的磷酸化、G蛋白相互作用等信号转导事件有关,RhSep1可能存在复杂的蛋白水平调控机制。