Improved sensorimotor adaptation after exhaustive exercise is accompanied by altered brain activity

Acute exercise has been shown to exhibit different effects on human sensorimotor behavior; however, the causes and mechanisms of the responses are often not clear. The primary aim of the present study was to determine the effects of incremental running until exhaustion on sensorimotor performance and adaptation in a tracking task. Subjects were randomly assigned to a running group (RG), a tracking group (TG), or a running followed by tracking group (RTG), with 10 subjects assigned to each group. Treadmill running velocity was initially set at 2.0 m s− 1, increasing by 0.5 m s− 1 every 5 min until exhaustion. Tracking consisted of 35 episodes (each 40 s) where the subjects' task was to track a visual target on a computer screen while the visual feedback was veridical (performance) or left-right reversed (adaptation). Resting electroencephalographic (EEG) activity was recorded before and after each experimental condition (running, tracking, rest). Tracking performance and the final amount of adaptation did not differ between groups. However, task adaptation was significantly faster in RTG compared to TG. In addition, increased alpha and beta power were observed following tracking in TG but not RTG although exhaustive running failed to induce significant changes in these frequency bands. Our results suggest that exhaustive running can facilitate adaptation processes in a manual tracking task. Attenuated cortical activation following tracking in the exercise condition was interpreted to indicate cortical efficiency and exercise-induced facilitation of selective central processes during actual task demands.

Analysis of rural activity spaces and transport disadvantage using a multi-method approach

Current knowledge about the relationship between transport disadvantage and activity space size is limited to urban areas, and as a result, very little is known about this link in a rural context. In addition, although research has identified transport disadvantaged groups based on their size of activity space, these studies have, however, not empirically explained such differences and the result is often a poor identification of the problems facing disadvantaged groups. Research has shown that transport disadvantage varies over time. The static nature of analysis using the activity space concept in previous research studies has lacked the ability to identify transport disadvantage in time. Activity space is a dynamic concept; and therefore possesses a great potential in capturing temporal variations in behaviour and access opportunities. This research derives measures of the size and fullness of activity spaces for 157 individuals for weekdays, weekends, and for a week using weekly activity-travel diary data from three case study areas located in rural Northern Ireland. Four focus groups were also conducted in order to triangulate quantitative findings and to explain the differences between different socio-spatial groups. The findings of this research show that despite having a smaller sized activity space, individuals were not disadvantaged because they were able to access their required activities locally. Car-ownership was found to be an important life line in rural areas. Temporal disaggregation of the data reveals that this is true only on weekends due to a lack of public transport services. In addition, despite activity spaces being at a similar size, the fullness of activity spaces of low-income individuals was found to be significantly lower compared to their high-income counterparts. Focus group data shows that financial constraint, poor connections both between public transport services and between transport routes and opportunities forced individuals to participate in activities located along the main transport corridors.

Visual voice activity detection using frontal versus profile views

Visual activity detection of lip movements can be used to overcome the poor performance of voice activity detection based solely in the audio domain, particularly in noisy acoustic conditions. However, most of the research conducted in visual voice activity detection (VVAD) has neglected addressing variabilities in the visual domain such as viewpoint variation. In this paper we investigate the effectiveness of the visual information from the speaker’s frontal and profile views (i.e left and right side views) for the task of VVAD. As far as we are aware, our work constitutes the first real attempt to study this problem. We describe our visual front end approach and the Gaussian mixture model (GMM) based VVAD framework, and report the experimental results using the freely available CUAVE database. The experimental results show that VVAD is indeed possible from profile views and we give a quantitative comparison of VVAD based on frontal and profile views The results presented are useful in the development of multi-modal Human Machine Interaction (HMI) using a single camera, where the speaker’s face may not always be frontal.

Which older women could benefit from interventions to decrease sitting time and increase physical activity?

In addition to the well-known health risks associated with lack of physical activity (PA), evidence is emerging about the health risks of sedentary behaviour (sitting). Research about patterns and correlates of sitting and PA in older women is scarce. METHODS: Self-report data from 6,116 women aged 76-81 years were collected as part of the Australian Longitudinal Study on Woman’s Health. Linear regression models were computed to examine whether demographic, social and health factors were associated with sitting and PA. RESULTS: Women who did no PA sat more than women who did any PA (p<0.001). Seven correlates were associated with sitting and PA (p<0.05). Five of these were associated with more sitting and less PA: three health-related (BMI, chronic conditions, anxiety/depression) and two social correlates (caring duties, volunteering). One demographic (being from another English-speaking country) and one social correlate (more social interaction) were associated with more sitting and more PA. Four correlates, two demographic (living in a city; post-high school education), one social (being single), and one health-related correlate (dizziness/loss of balance) were associated with more sitting only. Two other health-related correlates (stiff/painful joints; feet problems) were associated with less PA only. CONCLUSION: Sedentary behaviour and PA are distinct behaviours in older Australian women. Information about the correlates of both behaviours can be used to identify population groups who might benefit from interventions to reduce sedentary behaviour and/or increase PA.

Emergency health services : demand and service delivery models. Monograph 1: literature review and activity trends

Emergency Health Services (EHS), encompassing hospital-based Emergency Departments (ED) and pre-hospital ambulance services, are a significant and high profile component of Australia’s health care system and congestion of these, evidenced by physical overcrowding and prolonged waiting times, is causing considerable community and professional concern. This concern relates not only to Australia’s capacity to manage daily health emergencies but also the ability to respond to major incidents and disasters. EHS congestion is a result of the combined effects of increased demand for emergency care, increased complexity of acute health care, and blocked access to ongoing care (e.g. inpatient beds). Despite this conceptual understanding there is a lack of robust evidence to explain the factors driving increased demand, or how demand contributes to congestion, and therefore public policy responses have relied upon limited or unsound information. The Emergency Health Services Queensland (EHSQ) research program proposes to determine the factors influencing the growing demand for emergency health care and to establish options for alternative service provision that may safely meet patient’s needs. The EHSQ study is funded by the Australian Research Council (ARC) through its Linkage Program and is supported financially by the Queensland Ambulance Service (QAS). This monograph is part of a suite of publications based on the research findings that examines the existing literature, and current operational context. Literature was sourced using standard search approaches and a range of databases as well as a selection of articles cited in the reviewed literature. Public sources including the Australian Institute of Health and Welfare (AIHW), the Council of Ambulance Authorities (CAA) Annual Reports, Australian Bureau of Statistics (ABS) and Department of Health and Ageing (DoHA) were examined for trend data across Australia.

Structural and thermal performance of cold-formed steel stud wall systems under fire conditions

Cold-formed steel stud walls are a major component of Light Steel Framing (LSF) building systems used in commercial, industrial and residential buildings. In the conventional LSF stud wall systems, thin steel studs are protected from fire by placing one or two layers of plasterboard on both sides with or without cavity insulation. However, there is very limited data about the structural and thermal performance of stud wall systems while past research showed contradicting results, for example, about the benefits of cavity insulation. This research was therefore conducted to improve the knowledge and understanding of the structural and thermal performance of cold-formed steel stud wall systems (both load bearing and non-load bearing) under fire conditions and to develop new improved stud wall systems including reliable and simple methods to predict their fire resistance rating. Full scale fire tests of cold-formed steel stud wall systems formed the basis of this research. This research proposed an innovative LSF stud wall system in which a composite panel made of two plasterboards with insulation between them was used to improve the fire rating. Hence fire tests included both conventional steel stud walls with and without the use of cavity insulation and the new composite panel system. A propane fired gas furnace was specially designed and constructed first. The furnace was designed to deliver heat in accordance with the standard time temperature curve as proposed by AS 1530.4 (SA, 2005). A compression loading frame capable of loading the individual studs of a full scale steel stud wall system was also designed and built for the load-bearing tests. Fire tests included comprehensive time-temperature measurements across the thickness and along the length of all the specimens using K type thermocouples. They also included the measurements of load-deformation characteristics of stud walls until failure. The first phase of fire tests included 15 small scale fire tests of gypsum plasterboards, and composite panels using different types of insulating material of varying thickness and density. Fire performance of single and multiple layers of gypsum plasterboards was assessed including the effect of interfaces between adjacent plasterboards on the thermal performance. Effects of insulations such as glass fibre, rock fibre and cellulose fibre were also determined while the tests provided important data relating to the temperature at which the fall off of external plasterboards occurred. In the second phase, nine small scale non-load bearing wall specimens were tested to investigate the thermal performance of conventional and innovative steel stud wall systems. Effects of single and multiple layers of plasterboards with and without vertical joints were investigated. The new composite panels were seen to offer greater thermal protection to the studs in comparison to the conventional panels. In the third phase of fire tests, nine full scale load bearing wall specimens were tested to study the thermal and structural performance of the load bearing wall assemblies. A full scale test was also conducted at ambient temperature. These tests showed that the use of cavity insulation led to inferior fire performance of walls, and provided good explanations and supporting research data to overcome the incorrect industry assumptions about cavity insulation. They demonstrated that the use of insulation externally in a composite panel enhanced the thermal and structural performance of stud walls and increased their fire resistance rating significantly. Hence this research recommends the use of the new composite panel system for cold-formed LSF walls. This research also included steady state tensile tests at ambient and elevated temperatures to address the lack of reliable mechanical properties for high grade cold-formed steels at elevated temperatures. Suitable predictive equations were developed for calculating the yield strength and elastic modulus at elevated temperatures. In summary, this research has developed comprehensive experimental thermal and structural performance data for both the conventional and the proposed non-load bearing and load bearing stud wall systems under fire conditions. Idealized hot flange temperature profiles have been developed for non-insulated, cavity insulated and externally insulated load bearing wall models along with suitable equations for predicting their failure times. A graphical method has also been proposed to predict the failure times (fire rating) of non-load bearing and load bearing walls under different load ratios. The results from this research are useful to both fire researchers and engineers working in this field. Most importantly, this research has significantly improved the knowledge and understanding of cold-formed LSF walls under fire conditions, and developed an innovative LSF wall system with increased fire rating. It has clearly demonstrated the detrimental effects of using cavity insulation, and has paved the way for Australian building industries to develop new wall panels with increased fire rating for commercial applications worldwide.

Nobody says no : student self-censorship in a collaborative knowledge building activity

This paper explores student self-censorship within an online learning environment. Self-censorship in group activity can be seen as a two-edged sword. While it can be advantageous that a student censor personal frustration and angst when working with others, if the self-censorship impacts on the cognitive contribution a student makes then this may significantly impact upon the overall quality of the group’s collective knowledge artefact. This paper reports on a study where it was found that students had self censored both their feelings and ideas as they collaboratively worked together.

The defence of joint illegal activity must be looked at in context

In Miller v Miller (2011) 85 ALJR 480; [2011] HCA 9 the High Court examined the complex issue of joint illegal activity. The issue before the court was whether a plaintiff who had engaged in an illegal activity with the defendant may claim damages in negligence. In its decision the court analysed the cases of Henwood v Municipal Tramways Trust (SA) (1938) 60 CLR 438, Smith v Jenkins (1970) 119 CLR 397, Jackson v Harrison (1978) 138 CLR 438 and Gala v Preston (1991) 172 CLR 243.

The activity of learning assistance

This paper uses cultural-historical activity to examine the nature of learning assistance, provided in university settings, to assist learners in coping with academic study. Alternative rival constructions of learning assistance, either as part of an overall university activity system, or as an activity system overlapping with other separate activity systems (e.g. library and faculty) within the university, are outlined. Data from a research study at one university are used to describe the object, tools and tensions in learning assistance; and, together with the history of the service, these are used to suggest that the current situation is one where the objects of the Learning Assistance Unit, the faculty and the library have separated from the object of a university.

Measured and perceived physical fitness, intention, and self-reported physical activity in adolescence

Objective: The aim of this study was to investigate the associations among measured physical fitness, perceived fitness, intention towards future physical activity and self-reported physical activity through junior high school years. Methods: Study participants included 122 Finnish students who were 13 years old during Grade 7. The sample was comprised of 80 girls and 42 boys from 3 junior high schools (Grades 7-9). During the autumn semester of Grade 7, students completed fitness tests and a questionnaire analyzing self-perception of their physical fitness. The questionnaire delivered at Grade 8 included intention towards future physical activity. At Grade 9 students’ self-reported physical activity levels. Results: Structural Equation Modeling revealed an indirect path from physical fitness to self-reported physical activity via perceived physical fitness and intention towards future physical activity. The model also demonstrated a correlation between perceived physical fitness and physical activity. Squared multiple correlations revealed that perceived physical fitness explained 33 % of the actual physical fitness. Conclusions: The results of this study highlight the role of physical and cognitive variables in the process of adoption of physical activity in adolescence.

Substance-related expectancies among men who have sex with men : development of psychometric tools to predict unprotected sexual activity

Significant research has demonstrated direct and indirect associations between substance use and sexual behaviour. Substance use is related to sexual risk-taking and HIV seroconversion among some substance-using MSM. It remains unclear what factors mediate or underlie this relationship, and which substances are associated with greater harm. Substance-related expectancies are hypothesised as potential mechanisms. A conceptual model based on social-cognitive theory was tested, which explores the role of demographic factors, substance use, substance-related expectancies and novelty-seeking personality characteristics in predicting unprotected anal intercourse (UAI) while under the influence, across four commonly used substance types. Phase 1, a qualitative study (N = 20), explored how MSM perceive the effects of substance use on their thoughts, feelings and behaviours, including sexual behaviours. Information was attained through discussion and interviews, resulting in the establishment of key themes. Results indicated MSM experience a wide range of reinforcing aspects associated with substance use. General and specific effects were evident across substance types, and were associated with sexual behaviour and sexual risk-taking. Phase 2 consisted of developing a comprehensive profile of substance-related expectancies for MSM (SEP-MSM) regarding alcohol, cannabis, amyl nitrite and stimulants that possessed sound psychometric properties and was appropriate for use among this group. A cross-sectional questionnaire with 249 participants recruited through gay community networks was used to validate these measures, and involved online data collection, participants rating expectancy items and subsequent factor analysis. Results indicated expectancies can be reliably assessed, and predicted substance use patterns. Phase 3 examined demographic factors, substance use, substance-related expectancies, and novelty-seeking traits among another community sample of MSM (N = 277) throughout Australia, in predicting UAI while under the influence. Using a cross-sectional design, participants were recruited through gay community networks and completed online questionnaires. The SEP-MSM, and associated substance use, predicted UAI. This research extends social-cognitive theory regarding sexual behaviour, and advances understanding of the role of expectancies associated with substance use and sexual risk-taking. Future applications of the SEP-MSM in health promotion, prevention, clinical interventions and research are likely to contribute to reducing harm associated with substance-using MSM (e.g., HIV transmission).

Clusterin facilitates COMMD1 and I-kB degradation to enhance NF-kB activity in prostate cancer cells

Secretory clusterin (sCLU) is a stress-activated, cytoprotective chaperone that confers broad-spectrum cancer treatment resistance, and its targeted inhibitor (OGX-011) is currently in phase II trials for prostate, lung, and breast cancer. However, the molecular mechanisms by which sCLU inhibits treatment-induced apoptosis in prostate cancer remain incompletely defined. We report that sCLU increases NF-κB nuclear translocation and transcriptional activity by serving as a ubiquitin-binding protein that enhances COMMD1 and I-κB proteasomal degradation by interacting with members of the SCF-βTrCP E3 ligase family. Knockdown of sCLU in prostate cancer cells stabilizes COMMD1 and I-κB, thereby sequestrating NF-κB in the cytoplasm and decreasing NF-κB transcriptional activity. Comparative microarray profiling of sCLU-overexpressing and sCLU-knockdown prostate cancer cells confirmed that the expression of many NF-κB–regulated genes positively correlates with sCLU levels. We propose that elevated levels of sCLU promote prostate cancer cell survival by facilitating degradation of COMMD1 and I-κB, thereby activating the canonical NF-κB pathway.

A rebirth of physical activity in parents : applying a theory-based approach

Physical inactivity is a serious concern both nationally and internationally. Despite the numerous benefits of performing regular physical activity, many individuals lead sedentary lifestyles. Of concern, though, is research showing that some population sub-groups are less likely to be active, such as parents of young children. Although there is a vast amount of research dedicated to understanding people.s physical activity-related behaviours, there is a paucity of research examining those factors that influence parental physical activity. More importantly, research applying theoretical models to understand physical activity decision-making among this at-risk population is limited. Given the current obesity epidemic, the decline in physical activity with parenthood, and the many social and health benefits associated with regular physical activity, it is important that adults with young children are sufficiently active. In light of the dearth of research examining parental physical activity and the scant research applying a theory-based approach to gain this understanding, the overarching aim of the current program of research was to adopt a mixed methods approach as well as use sound theoretical frameworks to understand the regular physical activity behaviour of mothers and fathers with young children. This program of research comprised of three distinct stages: a qualitative stage exploring individual, social, and psychological factors that influence parental regular physical activity (Stage 1); a quantitative stage identifying the important predictors of parental regular physical activity intentions and behaviour using sound theoretical frameworks and testing a single-item measure for assessing parental physical activity behaviour (Stage 2); and a qualitative stage exploring strategies for an intervention program aimed at increasing parental regular physical activity (Stage 3). As a thesis by publication, eight papers report the findings of this program of research; these papers are presented according to the distinct stages of investigation that guided this program of research. Stage One of the research program comprised a qualitative investigation using a focus group/interview methodology with parents of children younger than 5 years of age (N = 40; n = 21 mothers, n = 19 fathers) (Papers 1, 2, and 3). Drawing broadly on a social constructionist approach (Paper 1), thematic analytic methods revealed parents. understandings of physical activity (e.g., requires effort), patterns of physical activity-related behaviours (e.g., grab it when you can, declining physical activity habits), and how constructions of social role expectations might influence parents. physical activity decision making (e.g., creating an active family culture, guilt and selfishness). Drawing on the belief-based framework of the TPB (Paper 2), thematic content analytic methods revealed parents. commonly held beliefs about the advantages (e.g., improves parenting practices), disadvantages (e.g., interferes with commitments), barriers (e.g., time), and facilitators (e.g., social support) to performing regular physical activity. Parents. normative beliefs about social approval from important others or groups (e.g., spouse/partner) were also identified. Guided by theories of social support, Paper Three identified parents. perceptions about the specific social support dimensions that influence their physical activity decision making. Thematic content analysis identified instrumental (e.g., providing childcare, taking over chores), emotional (e.g., encouragement, companionship), and informational support (e.g., ideas and advice) as being important to the decision-making of parents in relation to their regular physical activity behaviour. The results revealed also that having support for being active is not straightforward (e.g., guilt-related issues inhibited the facilitative nature of social support for physical activity). Stage Two of the research program comprised a quantitative examination of parents. physical activity intentions and behaviour (Papers 4, 5, 6, and 7). Parents completed an extended TPB questionnaire at Time 1 (N = 580; n = 288 mothers, n = 292 fathers) and self-reported their physical activity at Time 2, 1 week later (N = 458; n = 252 mothers, n = 206 fathers). Paper Four revealed key behavioural (e.g., improving parenting practices), normative (e.g., people I exercise with), and control (e.g., lack of time) beliefs as significant independent predictors of parental physical activity. A test of the TPB augmented to include the constructs of self-determined motivation and planning was assessed in Paper Five. The findings revealed that the effect of self-determined motivation on intention was fully mediated by the TPB variables and the impact of intention on behaviour was partially mediated by the planning variables. Slight differences in the model.s motivational sequence between the sexes were also noted. Paper Six investigated, within a TPB framework, a range of social influences on parents. intentions to be active. For both sexes, attitude, perceived behavioural control, group norms, friend general support, and an active parent identity predicted intentions, with subjective norms and family support further predicting mothers. intentions and descriptive norms further predicting fathers. intentions. Finally, the measurement of parental physical activity was investigated in Paper Seven of Stage Two. The results showed that parents are at risk of low levels of physical activity, with the findings also revealing validation support for a brief single-item physical activity measure. Stage Three of the research program comprised a qualitative examination of parents. (N = 12; n = 6 mothers, n = 6 fathers) ideas for strategies that may be useful for developing and delivering an intervention program aimed at increasing parental physical activity (Paper 8). Parents revealed a range of strategies for what to include in a physical activity intervention designed for parents of young children. For example, parents identified persuasion and information type messages, problem-solving strategies that engage parents in generating a priority list of their lifestyle commitments, and behavioural modification techniques such as goal setting and incentives. Social intervention strategies (e.g., social comparison, counselling) and environmental approaches (e.g., community-based integrative parent/child programs) were also identified as was a skill-based strategy in helping parents generate a flexible life/family plan. Additionally, a range of strategies for how to best deliver a parental physical activity intervention was discussed. Taken as a whole, Paper Eight found that adopting a multifaceted approach in both the design and implementation of a resultant physical activity intervention may be useful in helping to increase parental physical activity. Overall, this program of research found support for parents as a unique group who hold both similar and distinctive perceptions about regular physical activity to the general adult population. Thus, these findings highlight the importance of targeting intervention strategies for parents of young children. Additionally, the findings suggest that it might also be useful to tailor some messages specifically to each sex. Effective promotion of physical activity in parents of young children is essential given the low rate of activity in this population. Results from this program of research highlight parents as an at-risk group for inactivity and provide an important first step in identifying the factors that influence both mothers. and fathers. physical activity decision making. These findings, in turn, provide a foundation on which to build effective intervention programs aimed at increasing parents. regular physical activity which is essential for ensuring the health and well-being of parents with young children.

Rational design of serine protease inhibitors

Proteases regulate a spectrum of diverse physiological processes, and dysregulation of proteolytic activity drives a plethora of pathological conditions. Understanding protease function is essential to appreciating many aspects of normal physiology and progression of disease. Consequently, development of potent and specific inhibitors of proteolytic enzymes is vital to provide tools for the dissection of protease function in biological systems and for the treatment of diseases linked to aberrant proteolytic activity. The studies in this thesis describe the rational design of potent inhibitors of three proteases that are implicated in disease development. Additionally, key features of the interaction of proteases and their cognate inhibitors or substrates are analysed and a series of rational inhibitor design principles are expounded and tested. Rational design of protease inhibitors relies on a comprehensive understanding of protease structure and biochemistry. Analysis of known protease cleavage sites in proteins and peptides is a commonly used source of such information. However, model peptide substrate and protein sequences have widely differing levels of backbone constraint and hence can adopt highly divergent structures when binding to a protease’s active site. This may result in identical sequences in peptides and proteins having different conformations and diverse spatial distribution of amino acid functionalities. Regardless of this, protein and peptide cleavage sites are often regarded as being equivalent. One of the key findings in the following studies is a definitive demonstration of the lack of equivalence between these two classes of substrate and invalidation of the common practice of using the sequences of model peptide substrates to predict cleavage of proteins in vivo. Another important feature for protease substrate recognition is subsite cooperativity. This type of cooperativity is commonly referred to as protease or substrate binding subsite cooperativity and is distinct from allosteric cooperativity, where binding of a molecule distant from the protease active site affects the binding affinity of a substrate. Subsite cooperativity may be intramolecular where neighbouring residues in substrates are interacting, affecting the scissile bond’s susceptibility to protease cleavage. Subsite cooperativity can also be intermolecular where a particular residue’s contribution to binding affinity changes depending on the identity of neighbouring amino acids. Although numerous studies have identified subsite cooperativity effects, these findings are frequently ignored in investigations probing subsite selectivity by screening against diverse combinatorial libraries of peptides (positional scanning synthetic combinatorial library; PS-SCL). This strategy for determining cleavage specificity relies on the averaged rates of hydrolysis for an uncharacterised ensemble of peptide sequences, as opposed to the defined rate of hydrolysis of a known specific substrate. Further, since PS-SCL screens probe the preference of the various protease subsites independently, this method is inherently unable to detect subsite cooperativity. However, mean hydrolysis rates from PS-SCL screens are often interpreted as being comparable to those produced by single peptide cleavages. Before this study no large systematic evaluation had been made to determine the level of correlation between protease selectivity as predicted by screening against a library of combinatorial peptides and cleavage of individual peptides. This subject is specifically explored in the studies described here. In order to establish whether PS-SCL screens could accurately determine the substrate preferences of proteases, a systematic comparison of data from PS-SCLs with libraries containing individually synthesised peptides (sparse matrix library; SML) was carried out. These SML libraries were designed to include all possible sequence combinations of the residues that were suggested to be preferred by a protease using the PS-SCL method. SML screening against the three serine proteases kallikrein 4 (KLK4), kallikrein 14 (KLK14) and plasmin revealed highly preferred peptide substrates that could not have been deduced by PS-SCL screening alone. Comparing protease subsite preference profiles from screens of the two types of peptide libraries showed that the most preferred substrates were not detected by PS SCL screening as a consequence of intermolecular cooperativity being negated by the very nature of PS SCL screening. Sequences that are highly favoured as result of intermolecular cooperativity achieve optimal protease subsite occupancy, and thereby interact with very specific determinants of the protease. Identifying these substrate sequences is important since they may be used to produce potent and selective inhibitors of protolytic enzymes. This study found that highly favoured substrate sequences that relied on intermolecular cooperativity allowed for the production of potent inhibitors of KLK4, KLK14 and plasmin. Peptide aldehydes based on preferred plasmin sequences produced high affinity transition state analogue inhibitors for this protease. The most potent of these maintained specificity over plasma kallikrein (known to have a very similar substrate preference to plasmin). Furthermore, the efficiency of this inhibitor in blocking fibrinolysis in vitro was comparable to aprotinin, which previously saw clinical use to reduce perioperative bleeding. One substrate sequence particularly favoured by KLK4 was substituted into the 14 amino acid, circular sunflower trypsin inhibitor (SFTI). This resulted in a highly potent and selective inhibitor (SFTI-FCQR) which attenuated protease activated receptor signalling by KLK4 in vitro. Moreover, SFTI-FCQR and paclitaxel synergistically reduced growth of ovarian cancer cells in vitro, making this inhibitor a lead compound for further therapeutic development. Similar incorporation of a preferred KLK14 amino acid sequence into the SFTI scaffold produced a potent inhibitor for this protease. However, the conformationally constrained SFTI backbone enforced a different intramolecular cooperativity, which masked a KLK14 specific determinant. As a consequence, the level of selectivity achievable was lower than that found for the KLK4 inhibitor. Standard mechanism inhibitors such as SFTI rely on a stable acyl-enzyme intermediate for high affinity binding. This is achieved by a conformationally constrained canonical binding loop that allows for reformation of the scissile peptide bond after cleavage. Amino acid substitutions within the inhibitor to target a particular protease may compromise structural determinants that support the rigidity of the binding loop and thereby prevent the engineered inhibitor reaching its full potential. An in silico analysis was carried out to examine the potential for further improvements to the potency and selectivity of the SFTI-based KLK4 and KLK14 inhibitors. Molecular dynamics simulations suggested that the substitutions within SFTI required to target KLK4 and KLK14 had compromised the intramolecular hydrogen bond network of the inhibitor and caused a concomitant loss of binding loop stability. Furthermore in silico amino acid substitution revealed a consistent correlation between a higher frequency of formation and the number of internal hydrogen bonds of SFTI-variants and lower inhibition constants. These predictions allowed for the production of second generation inhibitors with enhanced binding affinity toward both targets and highlight the importance of considering intramolecular cooperativity effects when engineering proteins or circular peptides to target proteases. The findings from this study show that although PS-SCLs are a useful tool for high throughput screening of approximate protease preference, later refinement by SML screening is needed to reveal optimal subsite occupancy due to cooperativity in substrate recognition. This investigation has also demonstrated the importance of maintaining structural determinants of backbone constraint and conformation when engineering standard mechanism inhibitors for new targets. Combined these results show that backbone conformation and amino acid cooperativity have more prominent roles than previously appreciated in determining substrate/inhibitor specificity and binding affinity. The three key inhibitors designed during this investigation are now being developed as lead compounds for cancer chemotherapy, control of fibrinolysis and cosmeceutical applications. These compounds form the basis of a portfolio of intellectual property which will be further developed in the coming years.