从固体粗酶中提取α—淀粉酶的研究

本文以α－淀粉酶为研究对象，探索从固体粗酶浸取液中泡载浓缩α－淀粉酶的工艺过程，应用物料守恒原理对泡载分离过程进行了分析，实验分别考察了在浸取及泡载分离过程中酶活性的分离，提出了最优操作条件，并对酶的失活原因进行了探讨。     

物理法消除脱脂液泡沫机理分析

基于脱脂液中泡沫的物理化学性质,本文对离心消泡法、喷淋消泡法、加热消泡法和超声波消泡法等四种常见的物理消泡法的消泡机理进行了详细分析。不仅使工程技术人员对各种物理消泡方法的消泡机理有了深入认识,同时对消泡装置的开发设计也有重要的指导意义。     

掺杂钨丝中高温强化模式探讨

本文用扫描电镜和透射电镜观察了掺杂钨丝中钾泡的形成，借助微扰动理论描述了形成的动力学、在实验中还观察到这种钾泡与位错、晶界的相互作用，提出了这一相互作用的可能模式，按照这些模式对其所引起的强化作用初步进行了定量计算。结果表明，掺杂钨丝的中高温强化模式可能有所不同。     

AKS掺杂粒子演变及其丝材性能研究

通过对两组不同掺杂量的高温钼（Mo-silicon-aluminum-potassium简称AKS）AKS-2和AKS-4从烧结到后续压力加工整个过程中显微组织、力学性能进行分析对比研究；对高温钼的强化机理即K泡的形成，K泡的强化机理，K泡对再结晶温度的影响以及掺杂量对后续加工的影响进行了分析讨论，得出了AKS-4比AKS-2抗拉强度高，而延伸率却呈先增大后减小的趋势；掺杂量高的AKS-4再结晶温度比AKS-2高；AKS-4的再结晶温度区间也比AKS-2长；AKS-4比AKS-2的后续压力加工难度大，出现的缺陷也多，并分析了缺陷产生的原因。     

冷轧带钢表面鼓泡缺陷分析

采用扫描电镜对冷轧带钢表面鼓泡处的横、纵断面,鼓泡的破裂面以及鼓泡处拉断表面的形貌进行了观察,并对微区成分作了分析.鼓泡主要是在钢坯皮下由保护渣形成了层状夹杂,导致冷轧原料板出现分层,从而在轧制过程中心部、表面的塑性变形不一致而形成.     

含钴黄铁矿在悬浮固体鼓泡塔中间歇生物浸出期间细菌数的估算及与机械搅拌反应器的比较

S．Foucher等研究比较了在常规机械搅拌生物反应器与悬浮固体鼓泡塔反应器中生物浸出动力学与细菌数的变化。为此目的，分别在20L搅拌反应器和26L鼓泡塔中进行了间歇试验，悬浮物是含钴黄铁矿。在固体质量分数为20％条件下，两个反应器中的钴浸出率相似。在鼓泡塔中，生物浸出速率     

本钢冷轧厂2#镀锌机组清洗段消泡系统改造

介绍了本钢冷轧厂2#镀锌机组清洗段及消泡系统构成、作用及原理。针对原消泡系统存在的问题,采用气、水雾消泡技术,对消泡系统进行了改造。机组投入运行后,取得了良好效果。     

中低碳钢氢鼓泡形成机理分析

对中低碳钢氢致开裂(HIC)试验氢鼓泡进行了解剖分析。结果表明:氢鼓泡中均发现有夹杂物,包括A类硫化锰类夹杂、B类氧化铝类夹杂和大型的D类球形夹杂,所以氢鼓泡成因与A、B、D类夹杂均有关系。普通碳钢氢鼓泡中有片状硫化锰夹杂,而低硫钢氢鼓泡中主要是氧化铝类夹杂。氢鼓泡的大小与夹杂的大小和聚集程度有关。     

铁水脱硅与消泡机理及提高脱硅与消泡效率浅析

本文通过分析铁水脱硅与消泡热力学和动力学条件，从脱硅渣成分、碱度、粘度和表面张力等几方面研究了铁水脱硅与消泡机理，并结合太钢实际，探索了提高脱硅与消泡效率的途径。     

物理方法消除生物氧化过程中泡沫的探索研究

生物氧化过程中产生的大量泡沫不利于生产稳定运行,且易导致氧化效率降低、设备发生故障,采用化学方法消泡对生物氧化具有不利影响。通过进行物理方法消泡试验研究,如超声波、旋转叶片、离心力等消泡技术,探索一种较为切实可行的技术方案替代化学方法消泡。结果表明:物理方法相比化学方法消泡,具有生产成本低、消泡效率高等优势,且避免了消泡剂对菌种产生的不利影响,应用前景良好。离心力消泡技术已在辽宁天利金业有限责任公司成功实现工业化应用,消泡效果显著,泡沫层高度得到有效控制。     

pH- and Ca(2+)-dependent aggregation property of secretory vesicle matrix proteins and the potential role of chromogranins A and B in secretory vesicle biogenesis

Chromogranins A and B (CGA and CGB), the major proteins of the secretory vesicles of the regulated secretory pathway, have been shown to aggregate in a low pH and high calcium environment, the condition found in the trans-Golgi network where secretory vesicles are formed. Moreover, CGA and CGB, as well as several other secretory vesicle matrix proteins, have recently been shown to bind to the vesicle membrane at the intravesicular pH of 5.5 and to be released from it at a near physiological pH of 7.5. The pH- and Ca(2+)-dependent aggregation and interaction of chromogranins, as well as several other matrix proteins, with the vesicle membrane are considered essential in vesicle biogenesis. Therefore, to gain further insight into how vesicle matrix proteins find their way into the secretory vesicles, the pH- and Ca(2+)-dependent aggregation and vesicle membrane binding properties of the vesicle matrix proteins were studied, and it was found that most of the vesicle matrix proteins aggregated in the presence of Ca2+ at the intravesicular pH of 5.5. Furthermore, most of the vesicle matrix proteins bound not only to the vesicle membrane but also to CGA at pH 5.5, with the exception of a few matrix proteins that appeared to bind only to CGA or to vesicle membrane. Purified CGB was also shown to interact with CGA at pH 5.5. The extent and Ca(2+)-sensitivity of the aggregation of vesicle matrix proteins lay between those of purified CGB and CGA, CGB aggregation showing the highest degree of aggregation and being the most Ca2+ sensitive at a given protein concentration. Hence, in view of the abundance of chromogranins in secretory vesicles and their low pH- and high calcium-dependent aggregation property, combined with their ability to interact with both the vesicle matrix proteins and the vesicle membrane, CGA and CGB are proposed to play essential roles in the selective aggregation and sorting of potential vesicle matrix proteins to the immature secretory vesicles of the regulated secretory pathway.     

Studies on membrane fusion. I. Interactions of pure phospholipid membranes and the effect of myristic acid, lysolecithin, proteins and dimethylsulfoxide

The interaction and mixing of membrane components in sonicated unilamellar vesicles and also non-sonicated multilamellar vesicles prepared from highly purified phospholipids suspended in NaCl solutions has been examined. Electron microscopy and differential scanning calorimetry were used to characterize the extent and kinetics of mixing of membrane components between different vesicle populations. No appreciable fusion was detected between populations of non-sonicated phospholipid vesicles incubated in aqueous salt (NaCl) solutions. Mixing of vesicle membrane components via diffusion of phospholipid molecules between vesicles was observed in populations of negatively charged phosphatidylglycerol vesicles but similar exchange diffusion was not detected in populations of neutral phosphatidylcholine vesicles. Incubation of sonicated vesicle populations at temperatures close to or above the phospholipid transition temperature resulted in an increase in vesicle size and mixing of vesicle membrane components as determined by a gradual change in the thermotropic properties of the mixed vesicle population. The interaction of purified phospholipid vesicles was also examined in the presence of myristic acid and lysolecithin. Our results indicate that while these agents enhance mixing of vesicle membrane components, in most cases mixing probably proceeds via diffusion of phospholipid molecules rather than by fusion of entire vesicles. Increased mixing of vesicle membrane components was also produced when vesicles were prepared containing a purified hydrophobic protein (myelin proteolipid apoprotein) or were incubated in the presence of dimethylsulfoxide. In these two systems, however, the evidence suggests that mixing of membrane components results from the fusion of entire vesicles.     

A product of the Drosophila stoned locus regulates neurotransmitter release

The Drosophila stoned locus encodes two novel gene products termed stonedA and stonedB, which possess sequence motifs shared by proteins involved in intracellular vesicle traffic. A specific requirement for stoned in the synaptic vesicle cycle has been suggested by synthetic genetic interactions between stoned and shibire, a gene essential for synaptic vesicle recycling (Petrovich et al., 1993). A synaptic role of stoned gene products also is suggested by altered synaptic transients in electroretinograms recorded from stoned mutant eyes (Petrovich et al., 1993). We show here that the stonedA protein is highly enriched at Drosophila nerve terminals. Mutant alleles that affect stonedA disrupt the normal regulation of synaptic vesicle exocytosis at neuromuscular synapses of Drosophila. Spontaneous neurotransmitter release is enhanced dramatically, and evoked release is reduced substantially in such stoned mutants. Ultrastructural studies reveal no evidence of major disorganization at stoned mutant nerve terminals. Thus, our data indicate a direct role for stonedA in regulating synaptic vesicle exocytosis. However, genetic and morphological observations suggest additional, subtle effects of stoned mutations on synaptic vesicle recycling. Remarkably, almost all phenotypes of stoned mutants are similar to those previously described for mutants of synaptotagmin, a protein postulated to regulate both exocytosis and the recycling of synaptic vesicles. We propose a model in which stonedA functions together with synaptotagmin to regulate synaptic vesicle cycling.     

Multielectron excitations in the L-subshell photoabsorption of xenon

We have previously shown that the gas-vesicle protein GvpC is present on the outer surface of the gas vesicle, can be reversibly removed and rebound to the surface, and increases the critical collapse pressure of the gas vesicle. The GvpC molecule, which contains five partially conserved repeats of 33 amino acids (33-RR) sandwiched between 18 N-terminal and 10 C-terminal amino acids, is present in a ratio of 1:25 with the GvpA molecule, which forms the ribs of the gas vesicle. By using recombinant techniques we have now made modified versions of GvpC that contain only the first two, three or four of the 33-amino-acid repeats. All of these proteins bind to and strengthen gas vesicles that have been stripped of their native GvpC. Recombinant proteins containing three or four repeats bind in amounts that give the same ratio of 33-RR:GvpA (i.e. 1:5) as the native protein, and they restore much of the strength of the gas vesicle; the protein containing only two repeats binds at a lower ratio (1:7.7), however, and restores less of the strength. Ancestral proteins with only two, three or four of the 33-amino-acid repeats would have been functional in strengthening the gas vesicle but the progressive increase in number of repeats would have provided strength with increased efficiency.     

Preparation and characterization of echogenic liposome as an ultrasound contrast agent: size-dependency and stabilizing effect of cholesterol on the echogenicity of gas-entrapping liposome

The liposome entrapping CO2 gas inside the vesicle, which is called the echogenic liposome, has been made and characterized in vitro as an ultrasound contrast agent. The small unilamellar vesicle (SUV), large unilamellar vesicle (LUV) and multilamellar vesicle (MLV) as echogenic liposomes were compared in their echogenic efficiency and stability, and the effect of size and acoustic property was tested. The acoustic reflectivity increased with the increase in size of the vesicle, largest for the gas filled MLV among the three liposome suspensions. The acoustic reflectivity obtained with the echogenic MLV was larger than that of the gas bubbles enclosed within a surfactant mixture. A half-lifetime of 39 min was observed for the MLV prepared from egg-yolk phosphatidylcholine liposomes. The duration of reflectivity was prolonged drastically to a half-lifetime of 866 min by incorporating cholesterol into the MLV, although the echogenicity was decreased by such incorporation. The stabilizing effect of cholesterol for the ordinary liposomal membrane was thus ascertained in the present case of the gas-entrapping liposome. Our findings encourage the future development of improved gas-entrapping liposomes for the clinical trials of ultrasound contrast agents.     

Coupled ER to Golgi transport reconstituted with purified cytosolic proteins

A cell-free vesicle fusion assay that reproduces a subreaction in transport of pro-alpha-factor from the ER to the Golgi complex has been used to fractionate yeast cytosol. Purified Sec18p, Uso1p, and LMA1 in the presence of ATP and GTP satisfies the requirement for cytosol in fusion of ER-derived vesicles with Golgi membranes. Although these purified factors are sufficient for vesicle docking and fusion, overall ER to Golgi transport in yeast semi-intact cells depends on COPII proteins (components of a membrane coat that drive vesicle budding from the ER). Thus, membrane fusion is coupled to vesicle formation in ER to Golgi transport even in the presence of saturating levels of purified fusion factors. Manipulation of the semi-intact cell assay is used to distinguish freely diffusible ER- derived vesicles containing pro-alpha-factor from docked vesicles and from fused vesicles. Uso1p mediates vesicle docking and produces a dilution resistant intermediate. Sec18p and LMA1 are not required for the docking phase, but are required for efficient fusion of ER- derived vesicles with the Golgi complex. Surprisingly, elevated levels of Sec23p complex (a subunit of the COPII coat) prevent vesicle fusion in a reversible manner, but do not interfere with vesicle docking. Ordering experiments using the dilution resistant intermediate and reversible Sec23p complex inhibition indicate Sec18p action is required before LMA1 function.     

Uteroglobin-like antigen in the male genital tract secretions

The secretion of the rabbit seminal vesicle has been investigated by morphological and biochemical means. Apical regions of seminal vesicle epithelial cells show highly active protein synthesizing and releasing organells. The secretory proteins released are analysed by disc-electrophoresis and three different immunological procedures. There is evidence for the presence of an uteroglobin-like antigen in seminal vesicle secretion. Comparison with seminal plasma indicated that the uteroglobin-like protein is also present in this fluid. The immunological and electrophoretical identity of rabbit uteroglobin, obtained from the uterus, with "male uteroglobin" is obvious, but molecular-biochemical and biological identity awaits further clarification. The demonstration of uteroglobin-like antigen in the male as in the female points towards new aspects in reproductive and contraceptive research.     

Interaction of wheat alpha-thionin with large unilamellar vesicles

The interaction of the wheat antibacterial peptide alpha-thionin with large unilamellar vesicles has been investigated by means of fluorescence spectroscopy. Binding of the peptide to the vesicles is followed by the release of vesicle contents, vesicle aggregation, and lipid mixing. Vesicle fusion, i.e., mixing of the aqueous contents, was not observed. Peptide binding is governed by electrostatic interactions and shows no cooperativity. The amphipatic nature of wheat alpha-thionin seems to destabilize the membrane bilayer and trigger the aggregation of the vesicles and lipid mixing. The presence of distearoylphosphatidylethanolamine-poly(ethylene glycol 2000) (PEG-PE) within the membrane provides a steric barrier that inhibits vesicle aggregation and lipid mixing but does not prevent leakage. Vesicle leakage through discrete membrane channels is unlikely, because the release of encapsulated large fluorescent dextrans is very similar to that of 8-aminonaphthalene-1,3,6,trisulfonic acid (ANTS). A minimum number of 700 peptide molecules must bind to each vesicle to produce complete leakage, which suggests a mechanism in which the overall destabilization of the membrane is due to the formation of transient pores rather than discrete channels.     

The DnaJ-like cysteine string protein and exocytotic neurotransmitter release

The fast, tightly regulated release of neurotransmitters from presynaptic nerve terminals is effected by a complex molecular apparatus. The precise roles of the various proteins involved remain largely conjectural. Cysteine string proteins (CSPs) are novel synaptic vesicle components that have been conserved in evolution. They are characterized by an N-terminus 'J'-domain and a central, multiply palmitoylated string of cysteine residues. Vertebrate CSPs have been implicated in a functional interaction of synaptic vesicles with presynaptic Ca2+ channels. Genetic 'knockout' of CSPs in Drosophila results in a temperature-sensitive breakdown of elicited transmitter release. Here we try to integrate these observations into speculative functional models on the role of this new protein family in synaptic vesicle exocytosis.     

ADP-ribosylation factor-1 stimulates formation of nascent secretory vesicles from the trans-Golgi network of endocrine cells

ADP-ribosylation factor (ARF) is a small GTP-binding protein that has been implicated in intracellular vesicular transport. ARF regulates the budding of vesicles that mediate endoplasmic reticulum to Golgi and intra-Golgi transport. It also plays an important role in maintaining the function and morphology of the Golgi apparatus. Using a permeabilized cell system derived from GH3 cells, we provide evidence that ARF-1 regulates the formation of nascent secretory vesicles from the trans-Golgi network. Both myristoylated and non-myristoylated forms of recombinant human ARF-1 enhanced secretory vesicle budding about 2-fold. A mutant lacking the first 17 N-terminal residues, as well as one that preferentially binds GDP (T31N) did not stimulate vesicle formation. In contrast, a mutant defective in GTP hydrolysis (Q71L) promoted vesicle budding. Strikingly, a peptide corresponding to the N terminus of human ARF-1 (amino acids 2-17) also stimulated vesicle budding from the trans-Golgi network, in marked contrast to its inhibitory effect on vesicular transport from the endoplasmic reticulum to Golgi. These data demonstrate that in endocrine cells, ARF-1 and in particular its N terminus play an essential role in the formation of secretory vesicles.