Protein complementation assay as a display system for screening protein libraries in the intracellular environment

A wide range of screening strategies have been employed to isolate antibodies and other proteins with specific attributes, including binding affinity, specificity, stability and improved expression. However, there remains no high-throughput system to screen for target-binding proteins in a mammalian, intracellular environment. Such a system would allow binding reagents to be isolated against intracellular clinical targets such as cell signalling proteins associated with tumour formation (p53, ras, cyclin E), proteins associated with neurodegenerative disorders (huntingtin, betaamyloid precursor protein), and various proteins crucial to viral replication (e.g. HIV-1 proteins such as Tat, Rev and Vif-1), which are difficult to screen by phage, ribosome or cell-surface display. This study used the β-lactamase protein complementation assay (PCA) as the display and selection component of a system for screening a protein library in the cytoplasm of HEK 293T cells. The colicin E7 (ColE7) and Immunity protein 7 (Imm7) *Escherichia coli* proteins were used as model interaction partners for developing the system. These proteins drove effective β-lactamase complementation, resulting in a signal-to-noise ratio (9:1 – 13:1) comparable to that of other β-lactamase PCAs described in the literature. The model Imm7-ColE7 interaction was then used to validate protocols for library screening. Single positive cells that harboured the Imm7 and ColE7 binding partners were identified and isolated using flow cytometric cell sorting in combination with the fluorescent β-lactamase substrate, CCF2/AM. A single-cell PCR was then used to amplify the Imm7 coding sequence directly from each sorted cell. With the screening system validated, it was then used to screen a protein library based the Imm7 scaffold against a proof-of-principle target. The wild-type Imm7 sequence, as well as mutants with wild-type residues in the ColE7- binding loop were enriched from the library after a single round of selection, which is consistent with other eukaryotic screening systems such as yeast and mammalian cell-surface display. In summary, this thesis describes a new technology for screening protein libraries in a mammalian, intracellular environment. This system has the potential to complement existing screening technologies by allowing access to intracellular proteins and expanding the range of targets available to the pharmaceutical industry.

Self excitation in equity indices

A "self-exciting" market is one in which the probability of observing a crash increases in response to the occurrence of a crash. It essentially describes cases where the initial crash serves to weaken the system to some extent, making subsequent crashes more likely. This thesis investigates if equity markets possess this property. A self-exciting extension of the well-known jump-based Bates (1996) model is used as the workhorse model for this thesis, and a particle-filtering algorithm is used to facilitate estimation by means of maximum likelihood. The estimation method is developed so that option prices are easily included in the dataset, leading to higher quality estimates. Equilibrium arguments are used to price the risks associated with the time-varying crash probability, and in turn to motivate a risk-neutral system for use in option pricing. The option pricing function for the model is obtained via the application of widely-used Fourier techniques. An application to S&P500 index returns and a panel of S&P500 index option prices reveals evidence of self excitation.

A dynamic navigation model for unmanned aircraft systems and an application to autonomous front-on environmental sensing and photography using low-cost sensor systems

This paper presents an unmanned aircraft system (UAS) that uses a probabilistic model for autonomous front-on environmental sensing or photography of a target. The system is based on low-cost and readily-available sensor systems in dynamic environments and with the general intent of improving the capabilities of dynamic waypoint-based navigation systems for a low-cost UAS. The behavioural dynamics of target movement for the design of a Kalman filter and Markov model-based prediction algorithm are included. Geometrical concepts and the Haversine formula are applied to the maximum likelihood case in order to make a prediction regarding a future state of a target, thus delivering a new waypoint for autonomous navigation. The results of the application to aerial filming with low-cost UAS are presented, achieving the desired goal of maintained front-on perspective without significant constraint to the route or pace of target movement.

Rejoinder to James Hamilton

Hamilton (2001) makes a number of comments on our paper (Harding and Pagan, 2002b). The objectives of this rejoinder are, firstly, to note the areas in which we agree; secondly, to define with greater clarity the areas in which we disagree; and, thirdly, to point to other papers, including a longer version of this response, where we have dealt with some of the issues that he raises. The core of our debate with him is whether one should use an algorithm with a specified set of rules for determining the turning points in economic activity or whether one should use a parametric model that features latent states. Hamilton begins his criticism by stating that there is a philosophical distinction between the two methods for dating cycles and concludes that the method we use “leaves vague and intuitive exactly what this algorithm is intended to measure”. Nothing is further from the truth. When seeking ways to decide on whether a turning point has occurred it is always useful to ask the question, what is a recession? Common usage suggests that it is a decline in the level of economic activity that lasts for some time. For this reason it has become standard to describe a recession as a decline in GDP that lasts for more than two quarters. Finding periods in which quarterly GDP declined for two periods is exactly what our approach does. What is vague about this?

James Street market, Brisbane : a subtropical retail case study

Climate change mitigation is driving demand for energy-efficient and environmentally conscious commercial buildings in Australia. In the Australian subtropics, high rainfall, warm weather and humidity present unique challenges and opportunities for the architects tasked with designing eco-sensitive projects. The case of the James Street Market in Brisbane’s Fortitude Valley shows that climate-responsive design is an effective approach for reducing the environmental impact of commercial developments. The James Street Market combines climate-responsiveness, environmentally sensitive design strategies and smart planning to create a more sustainable retail precinct. This paper details the design strategies featured in the James Street Market, the project that kicked off a renaissance in climate-responsive commercial building design in Brisbane.

Selling James Bond : product placement in the James Bond films

The character of James Bond for many people is intrinsically linked in their minds with particular brands – Aston Martin, Bollinger, Omega, Smirnoff vodka, and so on. This direct association between character and brand highlights the intrinsic role of product placement in the film industry, and in the James Bond films in particular. Selling James Bond: Product Placement in the James Bond Films provides a comprehensive overview of the history of product placement in the James Bond series – charting the progression of the practice and drawing direct correlations to significant cultural and historical events that impacted upon the number and types of products incorporated into the series. While primarily a financial arrangement, it is also important that the practice of product placement be examined and understood in relation to these cultural contexts, an area of research so far largely ignored by academic study. Through extensive content analysis of the official James Bond film series, as well as utilising directors’ commentary and industry reports, this book illustrates the strong impact specific cultural and historical events have had on the practice of product placement in the series. In doing so, it provides an exciting and in-depth “behind the scenes” look at the James Bond film series, and its complicated and sometimes contentious history of product placement. In the process, it charts the gradual emergence of product placement from the more traditional background shot to becoming so embedded in the actual film narrative that they have become simply yet another method for filmmakers to produce cultural meaning.

Implementing a systematic process for rapidly embedding sustainability within chemical engineering education: a case study of James Cook University, Australia

Sustainability has emerged as a primary context for engineering education in the 21st Century, particularly the sub-discipline of chemical engineering. However, there is confusion over how to go about integrating sustainability knowledge and skills systemically within bachelor degrees. This paper addresses this challenge, using a case study of an Australian chemical engineering degree to highlight important practical considerations for embedding sustainability at the core of the curriculum. The paper begins with context for considering a systematic process for rapid curriculum renewal. The authors then summarise a 2-year federally funded project, which comprised piloting a model for rapid curriculum renewal led by the chemical engineering staff. Model elements contributing to the renewal of this engineering degree and described in this paper include: industry outreach; staff professional development; attribute identification and alignment; program mapping; and curriculum and teaching resource development. Personal reflections on the progress and process of rapid curriculum renewal in sustainability by the authors and participating engineering staff will be presented as a means to discuss and identify methodological improvements, as well as highlight barriers to project implementation. It is hoped that this paper will provide an example of a formalised methodology on which program reform and curriculum renewal for sustainability can be built upon in other higher education institutions.

A boy and his toys : technology and gadgetry in the James Bond films

This chapter provides an indepth examination of the history of product placement in the James Bond film series, specifically focusing on the emergence of technology and gadgetry in the series and the impact this had on the number and types of products that were placed in the films.

Licensed to sell : product placement in the James Bond film series

As one of the longest running franchises in cinema history, and with its well-established use of product placements, the James Bond film series provides an ideal framework within which to measure and catalogue the number and types of products used within a particular timeframe. This case study will draw upon extensive content analysis of the James Bond film series in order to chart the evolution of product placement across the franchise's 50 year history.