Barry Ryan

Line up, line up: using technology to align and enhance peer learning and assessment in a student centred foundation organic chemistry module

This paper describes how three technologies were utilised in combination to align student learning and assessment as part of a case study. Multiple choice questions (MCQs) were central to all these technologies. The peer learning technologies; Personal Response Devices (a.k.a. Clickers) and PeerWise (http://peerwise.cs.auckland.ac.nz), were implemented to achieve scaffolded, self-directed independent learning by the students which aligned to the assessment methodology through creating, analysing, answering and discussing multiple choice questions. Personal response devices enhanced in-class activity involvement, whilst PeerWise provided structure and support for independent student learning through defined outside class activities. An associated technology, online MCQs hosted though a secure virtual learning environment, was used as an aligned assessment methodology. The rationale behind this case study, its implementation and evaluation are described and discussed. Finally, the potential widespread applicability of this aligned, technology enhanced learning and assessment methodology is outlined along with suggestions and guidelines to aid practitioners wishing to implement a similar approach.

A walk down the red carpet: students as producers of digital video-based knowledge

Disengaged and apathetic students are common in many undergraduate classrooms. One approach to engage these students, and enhance the learning experience, is to integrate active learning into the curriculum. The purpose of this pedagogical evaluative study was to investigate if student researched, designed and created digital video could act as viable reusable peer learning resources. Although the use and integration of technology was central to the scope of this project, other ideas such as threshold concepts, the requirement for both active and authentic social constructivist learning, and student empowerment are pivotal to the rationalisation of this research. Overall, students appreciated an alternative method of learning; however, they were more reserved about their perceived learning and the usefulness of the peer-generated videos as learning resources. This pedagogical evaluative study suggests that practitioners can introduce student produced digital media as an alternative student centred learning approach whilst simultaneously developing student soft skills.

Consensus mutagenesis reveals that non-helical regions influence thermal stability of horseradish peroxidase

The enzyme horseradish peroxidase has many uses in biotechnology but a stabilized derivative would have even wider applicability. To enhance thermal stability, we applied consensus mutagenesis (used successfully with other proteins) to recombinant horseradish peroxidase and generated five single-site mutants. Unexpectedly, these mutations had greater effects on steady-state kinetics than on thermal stability. Only two mutants (T102A, T110V) marginally exceeded the wild types thermal stability (4% and 10% gain in half-life at 50 degrees C respectively); the others (Q106R, Q107D, I180F) were less stable than wild type. Stability of a five-fold combination mutant matched that of Q106R, the least-stable single mutant. These results were perplexing: the Class III plant peroxidases display wide differences in thermal stability, yet the consensus mutations failed to reflect these natural variations. We examined the sequence content of Class III peroxidases to determine if there are identifiable molecular reasons for the stability differences observed. Bioinformatic analysis validated our choice of sites and mutations and generated an archetypal peroxidase sequence for comparison with extant sequences. It seems that both genetic variation and differences in protein stability are confined to non-helical regions due to the presence of a highly conserved alpha-helical structural scaffold in these enzymes.

Learning in the Science Lab: a New Approach

This project aimed to improve the laboratory learning experience for undergraduate science students, focusing initially on first and third year cohorts, through specific objectives. Firstly, to incorporate novel teaching and assessment methods, including student led laboratories, in-house produced instructional videos, „Clickers‟ audience response devices, and pre-practical on-line MCQ assessments. Secondly, to develop timely feedback mechanisms, including peer review, tutor face to face and audio feedback, online automatic feedback, and report checklists. Finally, to imbed transferable skills into the laboratory including group work, communication skills (written and oral), organisation & project planning, health & safety, and preparedness for laboratories, final year projects & placement. Pedagogical evaluation was through anonymous multiple choice questionnaires and independent academic facilitated discussion forums. The main benefits are students who are better prepared, both for basic undergraduate laboratories and for independent research-based final year projects; continuity in the development of transferable skills; improved assessment quality though constructive alignment and appropriate feedback; and improved student satisfaction through engagement and feedback. The key recommendations arising from this study are; to encourage preparedness for practical sessions, harnessing technology to engage students through interesting pre-practical activities; to encourage an improved culture of feedback, including mechanisms such as podcasts, which also „feedforward‟; and to encourage a culture where value is added to modules by actively incorporating transferable skills into all student activities and assessments, rather than a „bolt on‟ approach.

Graduate Teaching Assistants; Critical Colleagues or Casual Components in the Undergraduate Laboratory Learning? An Exploration of the Role of the Postgraduate Teacher in the Sciences.

Abstract : Laboratory training is key to many science subjects and those that teach the practical laboratory skills maintain a pivotal role in undergraduate science training. Graduate Teaching Assistants (GTAs) are regularly used in higher education institutes to teach these practical lab skills. The GTA can be involved in both laboratory teaching and assessing all levels of undergraduates. This varied and challenging role requires support from the institute and if appropriately provided the learning experience can be rewarding not only for the undergraduate, but also for the GTA. In this review, the critical role of the laboratory GTA will be examined, their support requirement highlighted and the multi-dimensional benefits of GTA facilitated research-orientated laboratory learning outlined.

Overview of Approaches to Preventing and Avoiding Proteolysis During Expression and Purification of Proteins

Proteases are enzymes that cleave proteins. They occur widely in nature and play a fundamental role in cellular homeostasis; however, their presence can result in unwanted protein degradation during recombinant protein expression and purification. This unit introduces proteases, specifically focusing on the types commonly encountered during production of recombinant proteins. The strategies used to avoid and to prevent proteolysis are also described, with extensive consideration of the molecular, technical, and logistic methodologies involved. Curr. Protoc. Protein Sci. 71:5.25.1‐5.25.7.

A short chain NAD(H)-dependent alcohol dehydrogenase (HpSCADH) from Helicobacter pylori: A role in growth under neutral and acidic conditions

Toxic aldehydes produced by alcohol dehydrogenases have been implicated in the pathogenesis of Helicobacter pylori-related damage to the gastric mucosa. Despite this, the enzymes that might be responsible for producing such aldehydes have not been fully described. It was, therefore, of considerable interest to characterize the alcohol oxidizing enzymes in this pathogen. Previous work in this laboratory characterized two such H. pylori enzymes that had broad specificity for a range of aromatic alcohol substrates. However, an enzyme with specificity for aliphatic alcohols is likely to be required in order that H. pylori can metabolize the wide range of substrates encountered in the gastric mucosa. In this study we describe HpSCADH, an alcohol dehydrogenase from H. pylori 26695 with broad specificity for aliphatic alcohols. HpSCADH was classified in the cD1e subfamily of classical short chain alcohol dehydrogenases. The enzyme was a monomer of approximately 29kDa with a preference for NAD(+) as cofactor. Pyrazole was found to be a competitive inhibitor of HpSCADH. The physiological role of this enzyme was explored by construction of an HpSCADH isogenic mutant. At pH 7.0 the mutant showed reduced growth which became more pronounced when the pH was lowered to 5.0. When pyrazole was added to wild type H. pylori cells it caused growth profiles to be reduced to match those of the isogenic mutant suggesting that HpSCADH inhibition alone was responsible for growth impairment. Taken together, the data relating to the alcohol metabolizing enzymes of this pathogen indicate that they play an important role in H. pylori growth and adaptation to acidic environments. The therapeutic potential of targeting H. pylori alcohol dehydrogenases is discussed.

Differential Precipitation and Solubilization of Proteins

Differential protein precipitation is a rapid and economical step in protein purification and is based on exploiting the inherent physico-chemical properties of the polypeptide. Precipitation of recombinant proteins, lysed from the host cell, is commonly used to concentrate the protein of choice before further polishing steps with more selective purification columns (e.g. His-Tag, Size Exclusion, etc.). Recombinant proteins can also precipitate naturally as inclusion bodies due to various influences during over-expression in the host cell. Although this phenomenon permits easier initial separation from native proteins, these inclusion bodies must carefully be differentially solubilised so as to reform functional, correctly folded proteins. Here, a typical protein extraction, precipitation, and selective resolubilisation procedure is outlined, based on a recombinantly expressed protein.

Avoiding Proteolysis During Protein Chromatography

All cells contain proteases, which effect catalytic hydrolysis of the peptide bond between amino acids in the protein backbone. Typically, proteinases are prevented from nonspecific proteolysis by regulation and physical separation into different subcellular compartments; however, this segregation is not retained during cell lysis to release a protein of interest. Prevention of proteolysis during protein purification often takes the form of a two-pronged approach; first the inhibition of proteolysis in situ, followed by the separation of the protease from the protein of interest via chromatographical purification. Proteinase inhibitors are routinely used to limit the effect of the proteinases before they are physically separated from the protein of interest via column chromatography. Here, commonly used approaches to reducing proteolysis during chromatography are reviewed.

New inhibitors of the Kvβ2 subunit from mammalian Kv1 potassium channels.

The role of the redox state of Kvβ subunits in the modulation of Kv1 potassium channels has been well documented over the past few years. It has been suggested that a molecule that binds to or inhibits the aldo-keto reductase activity of Kvβ might affect the modulation of channel properties. Previous studies of possible modulators of channel activity have shown that cortisone and some related compounds are able to physically dissociate the channel components by binding to a site at the interface between α and β subunits. Herein, we describe some new inhibitors of rat brain Kvβ2, identified using an assay based on multiple substrate turnover. This approach allows one to focus on molecules that specifically block NADPH oxidation. These studies showed that, at 0.5mM, 3,4-dihydroxphenylacetic acid (DOPAC) was an inhibitor of Kvβ2 turnover yielding a ∼ 40-50% reduction in the aldehyde reductase activity of this subunit. Other significant inhibitors include the bioflavinoid, rutin and the polyphenol resveratrol; some of the known cardioprotective effects of these molecules may be attributable to Kv1 channel modulation. Cortisone or catechol caused moderate inhibition of Kvβ2 turnover, and the aldo-keto reductases inhibitor valproate had an even smaller effect. Despite the importance of the Kv1 channels in a number of disease states, there have been few Kvβ2 inhibitors reported. While the ones identified in this study are only effective at high concentrations, they could serve as tools to decipher the role of Kvβ2 in vivo and, eventually, inform the development of novel therapeutics.