Biocatalytic Self‐Assembly of Nanostructured Peptide Microparticles using Droplet Microfluidics

Uniformly‐sized, nanostructured peptide microparticles are generated by exploiting the ability of enzymes to serve (i) as catalysts, to control self‐assembly within monodisperse, surfactant‐stabilized water‐in‐oil microdroplets, and (ii) as destabilizers of emulsion interfaces, to enable facile transfer of the produced microparticles to water. This approach combines the advantages of biocatalytic self‐assembly with the compartmentalization properties enabled by droplet microfluidics. Firstly, using microfluidic techniques, precursors of self‐assembling peptide derivatives and enzymes are mixed in the microdroplets which upon catalytic conversion undergo molecular self‐assembly into peptide particles, depending on the chemical nature of the precursors. Due to their amphiphilic nature, enzymes adsorb at the water‐surfactant‐oil interface of the droplets, inducing the transfer of peptide microparticles from the oil to the aqueous phase. Ultimately, through washing steps, enzymes can be removed from the microparticles which results in uniformely‐sized particles composed of nanostructured aromatic peptide amphiphiles.     

Stages of acquisition and cessation for adolescent smoking: an empirical integration

Adolescent cigarette smoking acquisition and cessation were integrated into a single nine-stages-of-change continuum using the transtheoretical model of change framework. Findings in a high school student sample (n > 700) showed that a few of the never smokers were planning to try smoking, and half of the current smokers were contemplating quitting. More than half of former smokers were long-term quitters. The high pros of smoking scores assessing coping benefits of cigarettes were related to smoking acquisition and the high con (disadvantages) scores to long-term abstinence. Never smokers were most tempted to try smoking when they anticipated that smoking would help reduce negative and increase positive mood. Current and former smokers were tempted due to peer cigarette offers and negative mood. These temptations were significantly reduced among ex-smokers.     

Nonpeptide cyclic cyanoguanidines as HIV-1 protease inhibitors: synthesis, structure-activity relationships, and X-ray crystal structure studies

Comparison of the high-resolution X-ray structures of the native HIV-1 protease and its complexes with the inhibitors suggested that the enzyme flaps are flexible. The movement at the tip of the flaps could be as large as 7 A. On the basis of this observation, cyclic cyanoguanidines have been designed, synthesized, and evaluated as HIV-1 protease (PR) inhibitors. Cyclic cyanoguanidines were found to be very potent inhibitors of HIV-1 protease. The choice of cyclic cyanoguanidines over cyclic guanidines was based on the reduced basicity of the former. X-ray structure studies of the HIV PR complex with cyclic cyanoguanidine demonstrated that in analogy to cyclic urea, cyclic cyanoguanidines also displace the unique structural water molecule. The structure-activity relationship of the cyclic cyanoguanidines is compared with that of the corresponding cyclic urea analogues. The differences in binding constants of the two series of compounds have been rationalized using high-resolution X-ray structure information.     

Bioactivation and irreversible binding of the cognition activator tacrine using human and rat liver microsomal preparations. Species difference

Tacrine's [1,2,3,4-tetrahydro-9-acridinamine monohydrochloride monohydrate, (THA)] metabolic fate was examined using human and rat liver microsomal preparations. Following 1-hr incubations with human microsomes, [14C]THA (0.4 microM) was extensively metabolized to 1-hydroxyTHA with trace amounts of 2-, 4-, and 7-hydroxyTHA also produced. Poor recovery of radioactivity in the postreaction incubates suggested association of THA-derived radioactivity with precipitated microsomal protein. After exhaustive extraction, 0.034, 0.145, 0.126, and 0.012 nmol eq bound/mg protein/60 min of THA-derived radioactivity was bound to human liver preparations H109, H111, H116, and H118, respectively. Preparations H109 and H118 were lower in P4501A2 content and catalytic activity as compared with preparations H111 and H116. Incubations of equimolar [14C]1-hydroxyTHA with human liver microsomes also resulted in binding to protein, although to a lesser extent than observed with THA. [14C]THA (0.4 microM) was incubated for 1 hr with rat liver microsomes (1 microM P-450) prepared from noninduced (N), phenobarbital (PB), isoniazid (I), and 3-methylcholanthrene (3-MC)-pretreated animals. In all incubations, 1-hydroxyTHA was the major biotransformation product detected. After exhaustive extraction, 0.048, 0.054, 0.049, and 0.153 nmol eq/mg protein/60 min of THA-derived radioactivity was bound to microsomal protein from N, PB, I, and 3-MC pretreated rats. Increased binding with 3-MC induced rat liver preparations suggests the involvement of the P-450 1A subfamily in THA bioactivation. Glutathione (5 mM) coincubation inhibited the irreversible binding of THA-derived radioactivity in both human and 3-MC-induced rat liver preparations, whereas human epoxide hydrase (100 micrograms/incubate) had a relative minor effect. A mechanism is proposed involving a putative quinone methide(s) intermediate in the bioactivation and irreversible binding of THA. A species difference in THA-derived irreversible binding exists between human and noninduced rat liver microsomes, suggesting that the rat is a poor model for studying the underlying mechanism(s) of THA-induced elevations in liver marker enzymes found in clinical investigations.     

Cysteine contributions to metal binding preference for Zn/Cd in the beta-domain of metallothionein

Previous studies showed that metals in the beta-domain of metallothionein(MT) are more readily exchangeable and the level of avidity is sitespecific. This is reflected by energy differences computed with a series ofsimulated structures derived from X-ray crystallography. In this study, weexamined further the contribution of each of the nine cysteines in thebeta-domain. By semi-empirical MNDO calculations, we observed that therelative average binding strength is the strongest for Cys21 to Cd[M4] andfor Cys26 to Zn[M3], except for the bridging cysteines. These resultssuggest that binding site preference for Zn/Cd is determined by bindingstrength between specific cysteines and metal ion species.     

Survey of domestic food handling practices in New Zealand

The purpose of this survey was to obtain information on the domestic meat and poultry handling practices of New Zealanders in order to support the development of quantitative risk models, as well as providing data to underpin food safety campaigns to consumers. A sample of 1000 New Zealand residents, over 18 years of age, were randomly selected from the electoral roll and asked to participate in a national postal food safety study during 2005. Three hundred and twenty six respondents completed and returned questionnaires containing usable answers, and most of these respondents 'always' prepared the main meal within the household. The majority of meat (84.6%) and poultry (62.9%) purchased by New Zealanders was fresh (rather than frozen), and most consumers (94.4%) claimed that the time taken from food selection to reaching their home was 1 h or less. The majority (approximately 64%) of fresh meat and poultry was frozen in the home and the most favoured method of thawing was at room temperature for up to 12 h. The most common time period for storing cooked or raw meat and poultry in domestic refrigerators was up to 2 days. Most survey respondents preferred their meat and poultry to be cooked either medium or well done. The most popular cooking method for chicken was roasting or baking, while most respondents preferred to pan-fry steak/beef cuts, minced beef or sausages/hamburgers. The potential for undercooking was greatest with pan-fried steak with 19.8% of respondents preferring to consume this meat raw or rare. In answer to questions relating to food handling hygiene practices, 52.2% of respondents selected a hand washing sequence that would help prevent cross contamination. However, it was estimated that 41% and 28% of respondents would use knives and kitchen surfaces respectively in a manner that could allow cross contamination. The data in this survey are self-reported and, particularly for the hygiene questions, respondents may report an answer that they perceive as being correct rather than reflecting their actual behaviour. Nevertheless, the data on food processing, transport, storage and cooking preferences represent useful inputs into the assessment of food safety along the meat and poultry food chains.