Measuring the interaction forces between protein inclusion bodies and an air bubble using an atomic force microscope

Interaction forces between protein inclusion bodies and an air bubble have been quantified using an atomic force microscope (AFM). The inclusion bodies were attached to the AFM tip by covalent bonds. Interaction forces measured in various buffer concentrations varied from 9.7 nN to 25.3 nN (+/- 4-11%) depending on pH. Hydrophobic forces provide a stronger contribution to overall interaction force than electrostatic double layer forces. It also appears that the ionic strength affects the interaction force in a complex way that cannot be directly predicted by DLVO theory. The effects of pH are significantly stronger for the inclusion body compared to the air bubble. This study provides fundamental information that will subsequently facilitate the rational design of flotation recovery system for inclusion bodies. It has also demonstrated the potential of AFM to facilitate the design of such processes from a practical viewpoint.

Developing a framework for assessment of the environmental determinants of walking and cycling

The focus for interventions and research on physical activity has moved away from vigorous activity to moderate-intensity activities, such as walking. In addition, a social ecological approach to physical activity research and practice is recommended. This approach considers the influence of the environment and policies on physical activity. Although there is limited empirical published evidence related to the features of the physical environment that influence physical activity, urban planning and transport agencies have developed policies and strategies that have the potential to influence whether people walk or cycle in their neighbourhood. This paper presents the development of a framework of the potential environmental influences on walking and cycling based on published evidence and policy literature, interviews with experts and a Delphi study. The framework includes four features: functional, safety, aesthetic and destination; as well as the hypothesised factors that contribute to each of these features of the environment. In addition, the Delphi experts determined the perceived relative importance of these factors. Based on these factors, a data collection tool will be developed and the frameworks will be tested through the collection of environmental information on neighbourhoods, where data on the walking and cycling patterns have been collected previously. Identifying the environmental factors that influence walking and cycling will allow the inclusion of a public health perspective as well as those of urban planning and transport in the design of built environments. (C) 2002 Elsevier Science Ltd., All rights reserved.

Combined in-fermenter extraction and cross-flow microfiltration for improved inclusion body processing

In this study we demonstrate a new in-fermenter chemical extraction procedure that degrades the cell wall of Escherichia coli and releases inclusion bodies (IBs) into the fermentation medium. We then prove that cross-flow microfiltration can be used to remove 91% of soluble contaminants from the released IBs. The extraction protocol, based on a combination of Triton X-100, EDTA, and intracellular T7 lysozyme, effectively released most of the intracellular soluble content without solubilising the IBs. Cross-flow microfiltration using a 0.2 mum ceramic membrane successfully recovered the granulocyte macrophagecolony stimulating factor (GM-CSF) IBs with removal of 91% of the soluble contaminants and virtually no loss of IBs to the permeate. The filtration efficiency, in terms of both flux and transmission, was significantly enhanced by infermenter Benzonase(R) digestion of nucleic acids following chemical extraction. Both the extraction and filtration methods exerted their efficacy directly on a crude fermentation broth, eliminating the need for cell recovery and re-suspension in buffer. The processes demonstrated here can all be performed using just a fermenter and a single cross-flow filtration unit, demonstrating a high level of process intensification. Furthermore, there is considerable scope to also use the microfiltration system to subsequently solubilise the IBs, to separate the denatured protein from cell debris, and to refold the protein using diafiltration. In this way refolded protein can potentially be obtained, in a relatively pure state, using only two unit operations. (C) 2004 Wiley Periodicals Inc.

Mediatizing educational policy: The journalistic field, science policy, and cross-field effects

This paper is concerned to demonstrate the usefulness of the theory of Bourdieu, including the concepts of field, logics of practice and habitus, to understanding relationships between media and policy, what Fairclough has called the 'mediatization' of policy. Specifically, the paper draws upon Bourdieu's accessible account of the journalistic field as outlined in On television and journalism. The usefulness of this work is illustrated through a case study of a recent Australian science policy, The chance to change. As this policy went through various iterations and media representations, its naming and structure became more aphoristic. This is the mediatization of contemporary policy, which often results in policy as sound bite. The case study also shows the cross-field effects of this policy in education, illustrating how today educational policy can be spawned from developments in other public policy fields.

Estimating the potential refolding yield of recombinant proteins expressed as inclusion bodies

Recombinant protein production in bacteria is efficient except that insoluble inclusion bodies form when some gene sequences are expressed. Such proteins must undergo renaturation, which is an inefficient process due to protein aggregation on dilution from concentrated denaturant. In this study, the protein-protein interactions of eight distinct inclusion-body proteins are quantified, in different solution conditions, by measurement of protein second virial coefficients (SVCs). Protein solubility is shown to decrease as the SVC is reduced (i.e., as protein interactions become more attractive). Plots of SVC versus denaturant concentration demonstrate two clear groupings of proteins: a more aggregative group and a group having higher SVC and better solubility. A correlation of the measured SVC with protein molecular weight and hydropathicity, that is able to predict which group each of the eight proteins falls into, is presented. The inclusion of additives known to inhibit aggregation during renaturation improves solubility and increases the SVC of both protein groups. Furthermore, an estimate of maximum refolding yield (or solubility) using high-performance liquid chromatography was obtained for each protein tested, under different environmental conditions, enabling a relationship between yield and SVC to be demonstrated. Combined, the results enable an approximate estimation of the maximum refolding yield that is attainable for each of the eight proteins examined, under a selected chemical environment. Although the correlations must be tested with a far larger set of protein sequences, this work represents a significant move beyond empirical approaches for optimizing renaturation conditions. The approach moves toward the ideal of predicting maximum refolding yield using simple bioinformatic metrics that can be estimated from the gene sequence. Such a capability could potentially screen, in silico, those sequences suitable for expression in bacteria from those that must be expressed in more complex hosts. (C) 2004 Wiley Periodicals, Inc.

Globalizing policy sociology in education: Working with Bourdieu

This paper uses Bourdieu to develop theorizing about policy processes in education and to extend the policy cycle approach in a time of globalization. Use is made of Bourdieu's concept of social field and the argument is sustained that in the context of globalization the field of educational policy has reduced autonomy, with enhanced cross-field effects in educational policy production, particularly from the fields of the economy and journalism. Given the social rather than geographical character of Bourdieu's concept of social fields, it is also argued that the concept can be, and indeed has to be, stretched beyond the nation to take account of the emergent global policy field in education. Utilizing Bourdieu's late work on the globalization of the economy through neo-liberal politics, we argue that a non-reified account of the emergent global educational policy field can be provided.

The effects of adolescent cannabis use: How should the empirical evidence inform drug policy for young people? : Meeting abstract

Ethics as a subject is now consistently taught in medical schools within Australia. The theoretical Ethical models used, and the associated clinical discussions, vary between schools. Registrars have further theoretical Ethics teaching within Psychiatry Fellowship Training, and ongoing clinical work that is likely to provide exposure to complex and frequent Ethical dilemmas. As Psychiatry Trainees approach subspecialty training in Child and Adolescent Psychiatry they therefore have a rich experience of both theoretical Ethics teaching and clinical exposure to Ethical issues. In this symposium, the difficulties Child and Adolescent Psychiatry Trainees may have in the integration of multiple theoretical Ethical models are discussed. It is suggested that these difficulties make Ethics Teaching for Child and Adolescent Psychiatry Trainees particularly challenging. This is important given the complex Ethical issues often present when working with Children and their Families. The three main Ethical models of Deontology, Virtue Ethics and Consequentialism are described and their usefulness for the Child and Adolescent Psychiatrist examined. Limitations of these models, and “Four Principles” approaches (such as that of Beauchamp and Childress), for Child and Adolescent Psychiatry, are also considered. Clinical cases are included for discussion. Finally, the ways in which these models may be used to enhance Child and Adolescent Psychiatry Training, and subsequent clinical practice as a Child and Adolescent Psychiatrist, are discussed. The integration of different theoretical Ethical models is considered, with implications identified for clinical practice.

The policy cycle and vernacular globalization: a case study of the creation of Vietnam National University - Hochiminh City

This article examines the policy cycle and vernacular globalization in the context of higher education reform in Vietnam. Through an analysis of the development of the Vietnam National University - Hochiminh City as part of the post-1986 reconstruction of Vietnamese higher education, the article considers the complex interrelationship between globalized policy discourses, national interests and history in Vietnam, and the specific politics of policy implementation within one institution. Vietnam National University - Hochiminh City was created through an amalgamation of a number of smaller universities, and against the backdrop of social and economic restructuring aimed at promoting industrialization and a market orientation within socialist governance. The article reveals the dynamic tension between these local and global influences on higher education policy and practice, and more specifically, the dilemmas associated with top-down policy implementation when a new organization consists of older organizations with powerful provenance and reputations. In so doing the article demonstrates the necessity to globalize policy theory.

Centrifugal processing of cell debris and inclusion bodies from recombinant Escherichia coli

The settling characteristics of cell debris and inclusion bodies prior to, and following, fractionation in a disc-stack centrifuge were measured using Cumulative Sedimentation Analysis (CSA) and Centrifugal Disc photosedimentation (CDS). The impact of centrifuge feedrate and repeated homogenisation on both cell debris and inclusion body collection efficiency was investigated. Increasing the normalised centrifuge feedrate (Q/Sigma) from 1.32 x 10(-9) m s(-1) to 3.97 x 10(-9) m s(-1) leads to a 36% increase in inclusion body paste purity. Purity may also be improved by repeated homogenisation. Increasing the number of homogeniser passes results in smaller cell debris size whilst leaves inclusion body size unaltered. At a normalised centrifuge feedrate of 2.65 x 10(-9) m s(-1), increasing the number of homogeniser passes from two (2) to ten (10) improved overall inclusion body paste purity by 58%. Grade-efficiency curves for both the cell debris and inclusion bodies have also been generated in this study. The data are described using an equation developed by Mannweiler (1989) with parameters of k = 0.15-0.26 and n = 2.5-2.6 for inclusion bodies, and k = 0.12-0.14 and n = 2.0-2.2 for cell debris. This is the first accurate experimentally-determined grade efficiency curve for cell debris. Previous studies have simply estimated debris grade efficiency curves using an approximate debris size distribution and grade efficiency curves determined with 'ideal particles' (e.g. spherical PVA particles). The findings of this study may be used to simulate and optimise the centrifugal fractionation of inclusion bodies from cell debris.

Strategic trade policy under uncertainty: Sufficient conditions for the optimality of ad valorem, specific and quadratic trade taxes

In most previous work on strategic trade policy the form of government intervention has been prescribed in advance. In this paper, we apply a solution concept discussed by Klemperer and Meyer for games in which the strategy space consists of the class of all (non state-contingent) price quantity schedules. We examine a series of specific assumptions on demand and supply conditions and derive the associated equilibrium trade policies. We derive welfare implications for all cases examined.