Production of Extracellular Acid Proteases by Aspergillus Clavatus

The production of extracellular acid proteases from Aspergillus clavatus was evaluated in a culture filtrate medium, with different carbon and nitrogen sources. The fungus was cultivated at three different temperatures during 10 days. The proteolytic activity was determined on haemoglobin pH 5.0 at 37 °C. The highest acid proteolytic activity (80 U/ml) was observed in culture medium containing glucose and gelatin at 1%(w/v) at 30 °C at the third day of incubation. Cultures developed in Vogel medium with glucose at 2%(w/v) showed at about 45% of proteolytic activity when compared to the cultures with 1% of the same sugar. The optimum pH of enzymatic activity was 2.0 and the enzyme was stable at pH values ranging from 2.0 to 4.0. The optimum temperature was 40 °C and the half-lives at 40, 45 and 50 °C were 30, 10 and 5 min, respectively. This revised version was published online in July 2006 with corrections to the Cover Date.     

2DE-MS中膜蛋白质组的研究进展

2-维凝胶电泳(2DE)具有高分辨率、高通量等特点,已被广泛地用于蛋白质组的研究.然而,2DE-MS在膜蛋白质组学研究方面却有其局限性,主要因为:膜蛋白具有低丰度、难溶、等电点时易沉淀、难酶解等特点.然而随着亚细胞分离技术和直接的生化方法富集等技术的发展,低丰度问题得到了极大的改善;增溶剂(尿素,硫脲),新的两性离子和非离子去垢剂,以及有机溶剂等的利用极大地改善了膜蛋白质组的溶解性能;同时,一些新的2DE技术的利用扩大了常规2DE的分离范围.在膜蛋白裂解方面,将酶解法与化学法(CNBr)相结合,另外先进的质谱技术的发展使得膜蛋白质组的研究在最近几年取得了较大的发展.现对2DE-MS途径中,膜的富集、膜蛋白的提取、分离、酶解、鉴定方面的进展进行综述.     

应用脂质体介导技术改变重组杆状病毒感染方法

昆虫重组杆状病毒表达系统是有效的真核表达系统之一,广泛应用于重组蛋白的生产。目前常采用将重组病毒直接注射入家蚕体内的方法进行感染表达,在实际操作过程中很容易造成病毒对环境的污染,具有潜在的危险性。为了严格控制作业环境重组病毒的扩散和潜在污染,开发安全、有效的感染方法显得非常必要。本研究直接将病毒基因组DNA导入家蚕体内取得同样的感染效果,探讨了利用阳离子脂质体介导下的避免病毒污染和提高感染效果的感染方法。     

Effects of post-teneral diet on foraging success of sterile male Mediterranean fruit flies

Post‐teneral diets containing protein have been shown to enhance the copulatory success of sterile male Mediterranean fruit flies, Ceratitis capitata (Wied.) (Diptera: Tephritidae). However, ingesting protein was also found to negatively affect male survival, in particular when males faced starvation following release in the field. Accordingly, the objective of the present study was to determine the effects of various post‐teneral diets, presented to sterile males prior to release, on their subsequent ability to forage for carbohydrates and protein in the field. Using mark‐release recapture and analytic biochemical methods, we found that both protein‐fed and protein‐deprived males foraged successfully for protein and sugar in a field enclosure when these resources were available. We conclude that protein‐fed sterile males are able to exploit sources of nutrition in the release environment, and their inability to overcome starvation is not a concern for control operations using the sterile insect technique.     

纤维素降解菌L-06的筛选、鉴定及其产酶条件的分析

从用于堆肥的水稻秸秆中初筛出一株高效纤维素降解菌L-06, 根据18S rRNA基因序列和菌株形态分析, 初步鉴定该菌为斜卧青霉(Penicillium decumbens)。研究了液体发酵培养基中氮源、碳源、表面活性剂、培养温度、起始pH以及接种量对该菌株各纤维素酶活力的影响。在最适条件下, 该菌的b-葡萄糖苷酶(BGL)、外切纤维素酶(CBH)于培养第3天酶活力分别达到1662 u/mL和2770 u/mL; 内切纤维素酶(EG)、滤纸糖化力(FPase)于培养第4天酶活力分别达到18064 u/mL和4035 u/mL。在产酶优化实验中, 该菌的EG和CBH在pH10的培养条件下分别保持了70%和43%的酶活性; 在50oC培养条件下EG和CBH分别保持了68%和59%的酶活性。表现出了耐热, 耐碱的特性。     

植物提取物微生物限量标准及检测技术进展

植物提取物的微生物检测是保证植物产品安全性的重要手段, 制定严格的植物提取物质量控制标准体系, 特别是功能性食品、食品添加剂和植物源日用化学品等产品中微生物的检测和控制, 对产品的质量及安全保证具有重要作用, 是影响植物提取业实现全面发展的关键问题。本文主要介绍了部分国家植物提取物的微生物限量标准和植物提取物微生物检测的国内外现状与发展趋势, 并就如何建立植物提取物微生物检测行业标准体系提出了若干建议。希望对我国植物提取业实现新时期跨越式发展提供参考。     

生物破乳剂产生菌的筛选及其方法研究

针对生物破乳剂产生菌筛选难的问题, 采用显色法、溶血细胞测试法、表面张力测定法和排油圈法从6种不同菌源对生物破乳菌产生菌进行了筛选。通过试验筛选得到了17株生物破乳剂产生菌, 其中24h内破乳率高于70%的破乳菌有5株; 油田含油污泥、采油废水生物处理污泥和污水处理厂剩余污泥是筛选破乳菌的较好的菌源; 显色法、溶血圈法存在检测范围的局限性; 表面张力测定法和排油圈法是最为简易和准确的生物表面活性剂产生菌的筛选方法, 采用模型乳状液对生物破乳剂产生菌进行筛选最为直接和准确, 但工作量大、所需时间长, 因此在筛选高效破乳菌时, 建议采用表面张力、排油圈法进行初筛, 而后通过模型乳状液破乳进行验证。     

Applications of dual-color fluorescence cross-correlation spectroscopy in antibody binding studies

Two-photon dual-color fluorescence cross-correlation spectroscopy (DC-FCCS) was applied to study the binding interactions of monoclonal antibodies (mAbs) and protein antigens. We measured the binding constant of the interaction of a 32-amino acid brain natriuretic peptide (BNP) with a mAbs and demonstrated the utility of DC-FCCS in studies of antibody sandwiches, trimolecular formations, where two different antibodies bind the same antigen simultaneously. We also show the use of DC-FCCS for monitoring competitive displacement of the labeled antibody in antibody-antigen complexes and subsequent determination of the pertinent dissociation rate (off-rate). The off-rate measurements were performed for two mAbs toward tissue inhibitor 1 of metalloproteinases (TIMP-1). From a methodological perspective, selection of the best labeling protocols and careful optimization of the FCCS instrumentation are essential to achieve the highest cross-correlation signal. When working in vitro, it is practical to generate a complete binding curve using the normalized cross-correlation signal and then fit the experimental points to a binding model. DC-FCCS offers the sensitivity and all other advantages of a solution phase fluorescence-based technique. For systems containing proteins of a similar size that interact without substantial changes in the fluorescence intensity, DC-FCCS serves as a preferred means of measuring solution phase binding constants.     

Modeling incomplete sterility in a sterile release program: interactions with other factors

Models were constructed for control of a pest species by the release of sterile insects and these models explored the consequences of incomplete sterility. This feature was then coupled with the lack of competitive ability of released insects, the immigration of insects from outside the control area, and the mode of population regulation (density independent vs. density dependent). Using the density-independent models, it was seen that the limits on residual fertility of treated insects become much more stringent when incomplete sterilization is combined with a lack of competitive ability and immigration of insects into the control area. Strong density dependence in the system has a marked moderating effect on the requirements for sterility, competitive ability, and immigration. However, if the density-independent limits on these factors are exceeded, then suppression is possible, but collapse of the pest population is impossible using sterile releases alone. Even suppression might not be satisfactory if these three detrimental factors are prominent. It is suggested that one remedy is the use of the sterile release method in combination with other control methods. Received: January 18, 2001 / Accepted: August 7, 2001     

黑曲霉SL2-111复合酶固体发酵工艺研究

以酸性蛋白酶酶活为响应值,采用单因素搜索和正交试验对黑曲霉（Aspergillus niger）SL2－111固体发酵工艺进行优化,结果表明最适培养基的组成为：新鲜麸皮8．25g,米糠4．5g、豆饼粉1．5g、（NH4)2SO40.3g、K2HPO40．66g、CaCl20.075g、水8．6mL,pH5．5,变温培养,前30h28℃、后30h为23℃,培养时间为60h。采用最适培养基和优化工艺,在250mL三角瓶中进行验证实验,酸性蛋白酶酶活可达12586U/g,果胶酶和纤维素酶分别为16490U/g、9822U／g。