Construction, properties and specific fluorescent labeling of a bovine prothrombin mutant engineered with a free C-terminal cysteine

To define the role of phosphatidylserine-induced conformationalchanges in prothrombin activation during blood coagulation,a recombinant bovine prothrombin was constructed, characterizedand shown to have a globally native-like conformation. We introduceda cysteine to replace the penultimate residue (Gly581) of apreviously constructed active site mutant, and expressed thedouble mutant in Chinese hamster ovary cells at the level of0.6 µg/ml of cell culture medium. Specific labeling withfluorescein maleimide was accomplished by limited reductionwith dithiothreitol to free the engineered cysteine while maintainingthe native-like functional properties of the molecule. The averagestoichiometry of labeling was 0.84 probe/protein. The locationof the probe at the C-terminus was confirmed by proteolysisby native thrombin, by Taipan venom, and by carboxypeptidaseY. Both the double mutant and labeled prothrombin could be activatedby snake venoms and the prothrombinase but, as expected, thedouble mutant meizothrombin did not autolyze as does nativemeizothrombin. Thus, for the first time, a native-like but specificallylabeled prothrombin has been constructed. This molecule willbe an essential tool for elucidating the structural role ofmembranes during prothrombin activation. In addition, the methodsdescribed might be usefully applied to labeling of an odd, engineeredcysteine in other disulfide bond-containing proteins.     

Structuration bridging diffusion of innovations and gender relations theories: a case of paradigmatic pluralism in IS research

This paper discusses the adoption of a pluralist theoretical framework – one that is also multiparadigmatic – for conducting and publishing information system (IS) research. The discussion is illustrated by a single case study involving the Australian cotton industry. The theoretical framework is informed by three sociological theories, each with its particular paradigmatic assumptions: structuration theory as a meta‐theory, and diffusion of innovations and gender relations as lower‐level theories from notionally opposing paradigms. Theoretical pluralism helped to produce rich findings, illuminating both the social nature of women farmers' roles, the materiality of the cotton farming context, the characteristics of the decision support systems in use and the recursive way in which human agency and institutional pressures shape each other. Because users of so‐called divergent paradigms often face criticism based on the incommensurability issue, one of the main contributions of this paper is to discuss the value of a pluralist and multiparadigmatic theoretical framework in dealing with complex IS social phenomena.     

Relationship between cell morphology and impact strength of microcellular foamed high‐density polyethylene/polypropylene blends

Polymer blends, such as those resulting from recycling postconsumer plastics, often have poor mechanical properties. Microcellular foams have been shown to have the potential to improve properties, and permit higher‐value uses of mixed polymer streams. In this study, the effects of microcellular batch processing conditions (foaming time and temperature) and HDPE/PP blend compositions on the cell morphology (the average cell size and cell‐population density) and impact strength were studied. Optical microscopy was used to investigate the miscibility and crystalline morphology of the HDPE/PP blends. Pure HDPE and PP did not foam well at any processing conditions. Blending facilitated the formation of microcellular structures in polyolefins because of the poorly bonded interfaces of immiscible HDPE/PP blends, which favored cell nucleation. The experimental results indicated that well‐developed microcellular structures are produced in HDPE/PP blends at ratios of 50:50 and 30:70. The cell morphology had a strong relationship with the impact strength of foamed samples. Improvement in impact strength was associated with well‐developed microcellular morphology. Polym. Eng. Sci. 44:1551–1560, 2004. © 2004 Society of Plastics Engineers.     

Family of G protein {alpha} chains: amphipathic analysis and predicted structure of functional domains

The G proteins transduce hormonal and other signals into regulationof enzymes such as adenylyl cyclase and retinal cGMP phosphodiesterase.Each G protein contains an subunit that binds and hydrolyzesguanine nucleotides and interacts with ß subunitsand specific receptor and effector proteins. Amphipathic andsecondary structure analysis of the primary sequences of fivedifferent chains (bovine _s, _t1 and _t2, mouse _i, and rat _o)predicted the secondary structure of a composite chain (_avg).The chains contain four short regions of sequence homologousto regions in the GDP binding domain of bacterial elongationfactor Tu (EF-Tu). Similarities between the predicted secondarystructures of these regions in _avg and the known secondary structureof EF-Tu allowed us to construct a three-dimensional model ofthe GDP binding domain of _avg. Identification of the GDP bindingdomain of _avg defined three additional domains in the compositepolypeptide. The first includes the amino terminal 41 residuesof _avg, with a predicted am phipathic helical structure; thisdomain may control binding of the chains to the ßcomplex. The second domain, containing predicted ßstrands and helices, several of which are strongly amphipathic,probably contains sequences responsible for interaction of chains with effector enzymes. The predicted structure of thethird domain, containing the carhoxy terminal 100 amino acids,is predominantly ß sheet with an amphipathic helixat the carboxy terminus. We propose that this domain is reponsiblefor receptor binding. Our model should help direct further experimentsinto the structure and function of the G protein chain.     

A comparison of pyrrolidide and picolinyl ester derivatives for the identification of fatty acids in natural samples by gas chromatography-mass spectrometry

The pyrrolidide and picolinyl ester derivatives of the fatty acids in two natural lipid samples rich in unsaturated fatty acids, pig testis lipids and cod liver oil were satisfactorily resolved on capillary columns of fused silica coated with stationary phases of varying polarity. The picolinyl esters, in particular, when subjected to gas chromatography-mass spectrometry on a column containing a cross-linked methyl silicone, gave distinctive mass spectra, which could be interpreted in terms of both the numbers and positions of the double bonds.     

An International Collaborative Study on Trypsin Inhibitor Assay for Legumes,Cereals, and Related Products

For determining trypsin inhibitor activity (TIA) in soy products, the American Oil Chemists' Society (AOCS) Method Ba 12-75 has been used. It measures differences in absorbance at 410 nm of bovine trypsin activity toward a synthetic substrate (Nα-benzoyl-DL-arginine-p-nitroanilide) in the absence and presence of an inhibitor. Recently, a significantly improved method was developed (JAOCS, 2019, 96:635–645), featuring 5 mL of total assay volume, enzyme-last sequence, and single inhibitor level in duplicate. It is proposed as the AOCS Method Ba 12a-2020. As a part of the AOCS method approval process, a collaborative study involving 12 international laboratories was conducted to evaluate the performance of the proposed method. The study involved measuring TIA in 10 selected test samples plus a blind duplicate. They included soybeans, pulses, cereals, and their processed products (flours, concentrates, and isolates). After rigorous statistical treatment of the data, only three outliers were removed from the data of two samples. Repeatability relative standard deviations (RSDr) for the 11 samples ranged from 0.99% to 5.52%. Reproducibility RSD (RSD_R) ranged from 7.07% to 22.92%, with seven samples having RSD_R around 10% or less. The remaining four samples had very low TIA, and their RSD_R values ranged from 13.34% to 22.92%. The study has demonstrated reliable performance of the proposed AOCS method. Several collaborators carried out additional experiments addressing some aspects of the method, leading to further refinements. The proposed method is undergoing evaluation by the AOCS Uniform Methods Committee for adoption as an Official Method for measuring TIA in various legume and grain products.     

用仿生自组装法制备的新型生物复合材料及表征

通过仿生自组装法在室温下制备了纳米碳酸化羟基磷灰石/胶原（nCHAC）复合材料,并采用X射线衍射（XRD）、热分析（TGA）和透射电镜（TEM）进行了观察和分析.结果表明,该材料具有与天然骨相同的纳米尺度和胶原蛋白无机相,含有（2.8～14.7）w%的碳酸化HA成分;其显微结构是矿化的胶原纤维束,类似于天然骨的分级结构.揭示了碳酸化HA和胶原含量不同的复合材料中,具有微小差别的自组装单元.碳酸的百分含量影响矿物的晶体尺寸和胶原纤维的组装.因此,nCHAC复合材料是有前途的硬组织修复用材料,其所具有的特殊性能要归功于生物仿生制备出的nCHAC材料有与天然骨类似的成分和类似的微观结构.     

Forming an intention to act on recommendations given via online social networks

Online social networks (OSN) are technology-enabled relationship tools in which a user creates a profile, connects to other individuals, and shares user-generated content with them. One of the many features OSN offer is the ability to post messages in the form of product and service recommendations. Although word-of-mouth research has examined this type of personal communication to intentionally influence consumer decisions, limited understanding exists regarding when a user acts upon a recommendation received from one of their contacts. In this study, we draw on relationship theories and research on trust to analyze the impact relationship characteristics and behaviors of the OSN contacts have on trust in the recommendation that subsequently leads a user to act on this OSN contact’s recommendation. The results of a survey of 116 OSN users showed that closeness, excessive posting behavior, and past recommendation experience have a positive impact on trust and intention to act on the recommendation. One characteristic of the relationships, that is, similarity, however, does not affect recommendation trust in the context of OSN. The findings enhance our understanding of relationships and their defining characteristics in OSN. The results also provide insights into how companies might leverage OSN in their marketing efforts.     

Awareness of Organizational Expertise

Comprehension of computer programs involves identifying important program parts and inferring relationships between them. The ability to comprehend a computer program is a skill that begins its development in the novice programmer and reaches maturity in the expert programmer. This research examined the beginning of this process, that of comprehension of computer programs by novice programmers. The mental representations of the program text that novices form, which indicate the comprehension strategies being used, were examined. In the first study, 80 novice programmers were tested on their comprehension of short program segments. The results suggested that novices form detailed, concrete mental representations of the program text, supporting work that has previously been done with novice comprehension. Their mental representations were primarily procedural in nature, with little or no modeling using real‐world referents. In a second study, the upper and lower quartile comprehenders from Study 1 were tested on their comprehension of a longer program. Results supported the conclusions from Study 1 in that the novices tended towards detailed representations of the program text with little real‐world reference. However, the comprehension strategies used by high comprehenders differed substantially from those used by low comprehenders. Results indicated that the more advanced novices were using more abstract concepts in their representations, although their abstractions were detailed in nature.