title: Understanding allosteric enzyme regulation using macromolecular rate theory

authors: Hamill, Carlin James

subjects: Enzyme

abstract: Enzyme catalysed reaction rates produce a curved temperature dependence with a temperature optimum, Topt. Traditionally enzyme temperature dependence has been modelled by the Arrhenius and Eyring-Polanyi equations. Deviations from
these models at elevated temperatures have traditionally been attributed to enzyme denaturation. Increasing evidence shows that enzyme denaturation is insufficient to fully explain the curvature for enzyme catalysed rates with temperature.

Macromolecular rate theory (MMRT) accounts for these deviations from early models by introducing the concept of heat capacity (Cp). During an enzymatic reaction, the heat capacity is reduced as the transition state complex forms (due to enzyme rigidification), resulting in an overall negative change in heat capacity (ΔCp^‡) for the reaction. Heat capacity is a function of the number and energies of the vibrational modes for the individual molecules. Enzyme-ligand binding will influence the energies of the vibrational modes of the molecular complex
therefore affecting an enzymes’ heat capacity and its catalytic rate via ΔCp^‡. Notably, a form of enzyme regulation (called allostery) utilises ligand binding at a site other than the active site to effect changes in catalytic rate.

The underlying mechanisms of allostery are poorly understood. Here, the mechanisms of allostery are explored using a model isomaltase enzyme and MMRT to investigate changes in heat capacity. Several mutants of the model MalL enzyme were designed, produced and purified. These mutations aimed to either
alter the structural integrity or mimic ligand binding in the C-terminal domain. Using a Stopped-Flow apparatus, these mutants were characterised kinetically and using crystallographic techniques, two structures were solved at high resolution.

Targeted mutations approximately 30 Å from the active site showed significant changes in enzyme rate. Overall, the two new structures of MalL were very similar to the apo-enzyme showing only very minor structural deviations. Changes to the rate can thus be attributed to changes in enzyme dynamics and heat capacity.
These results are a promising first step in linking changes in enzymatic activity with changes to enzyme dynamics and heat capacity, using MMRT as a model.

text: Understanding allosteric enzyme regulation using macromolecular rate theory Enzyme catalysed reaction rates produce a curved temperature dependence with a temperature optimum, Topt. Traditionally enzyme temperature dependence has been modelled by the Arrhenius and Eyring-Polanyi equations. Deviations from
these models at elevated temperatures have traditionally been attributed to enzyme denaturation. Increasing evidence shows that enzyme denaturation is insufficient to fully explain the curvature for enzyme catalysed rates with temperature.

Macromolecular rate theory (MMRT) accounts for these deviations from early models by introducing the concept of heat capacity (Cp). During an enzymatic reaction, the heat capacity is reduced as the transition state complex forms (due to enzyme rigidification), resulting in an overall negative change in heat capacity (ΔCp^‡) for the reaction. Heat capacity is a function of the number and energies of the vibrational modes for the individual molecules. Enzyme-ligand binding will influence the energies of the vibrational modes of the molecular complex
therefore affecting an enzymes’ heat capacity and its catalytic rate via ΔCp^‡. Notably, a form of enzyme regulation (called allostery) utilises ligand binding at a site other than the active site to effect changes in catalytic rate.

The underlying mechanisms of allostery are poorly understood. Here, the mechanisms of allostery are explored using a model isomaltase enzyme and MMRT to investigate changes in heat capacity. Several mutants of the model MalL enzyme were designed, produced and purified. These mutations aimed to either
alter the structural integrity or mimic ligand binding in the C-terminal domain. Using a Stopped-Flow apparatus, these mutants were characterised kinetically and using crystallographic techniques, two structures were solved at high resolution.

Targeted mutations approximately 30 Å from the active site showed significant changes in enzyme rate. Overall, the two new structures of MalL were very similar to the apo-enzyme showing only very minor structural deviations. Changes to the rate can thus be attributed to changes in enzyme dynamics and heat capacity.
These results are a promising first step in linking changes in enzymatic activity with changes to enzyme dynamics and heat capacity, using MMRT as a model.

year: 2020

title: Can we avoid high coupling?

authors: Taube-Schock, Craig

subjects: coupling

abstract: It is considered good software design practice to organize source code into modules and to favour within-module connections (cohesion) over between-module connections (coupling), leading to the oft-repeated maxim "low coupling/high cohesion". Prior research into network theory and its application to software systems has found evidence that many important properties in real software systems exhibit approximately scale-free structure, including coupling; researchers have claimed that such scale-free structures are ubiquitous. This implies that high coupling must be unavoidable, statistically speaking, apparently contradicting standard ideas about software structure. We present a model that leads to the simple predictions that approximately scale-free structures ought to arise both for between-module connectivity and overall connectivity, and not as the result of poor design or optimization shortcuts. These predictions are borne out by our large-scale empirical study. Hence we conclude that high coupling is not avoidable--and that this is in fact quite reasonable.

text: Can we avoid high coupling? It is considered good software design practice to organize source code into modules and to favour within-module connections (cohesion) over between-module connections (coupling), leading to the oft-repeated maxim "low coupling/high cohesion". Prior research into network theory and its application to software systems has found evidence that many important properties in real software systems exhibit approximately scale-free structure, including coupling; researchers have claimed that such scale-free structures are ubiquitous. This implies that high coupling must be unavoidable, statistically speaking, apparently contradicting standard ideas about software structure. We present a model that leads to the simple predictions that approximately scale-free structures ought to arise both for between-module connectivity and overall connectivity, and not as the result of poor design or optimization shortcuts. These predictions are borne out by our large-scale empirical study. Hence we conclude that high coupling is not avoidable--and that this is in fact quite reasonable.

year: 2011

title: Teachers' perceptions of the effectiveness of the Whānau Rūmaki teacher education programme, 1990-1992

authors: Kana, Fred

subjects: Maori Language

abstract: In 1990 the School of Education, the University of Waikato, initiated a special option within its regular pre-service teacher training programmes, an option designed to prepare teachers to deliver the New Zealand curriculum in bilingual and total immersion schools through the medium of te reo Māori. This alternative programmed, designated Whānau Rūmaki, was experimental and evolutionary. Inevitably particular elements of it had to be varied with each year's operation as its architects attempted 'to get the mix right'. It is difficult, however, when engaged in the day to day running of a programme, to stand back and review with any degree of objectivity its strengths and weaknesses. This study, therefore, in retrospect attempts to do three things: one, to follow the initial Whānau Rūmaki student cohort through its three or four year training cycle noting especially the difficulties confronting individual students some of whom dropped out of the programme; two, to gather information from as many as possible of the graduates who graduated about how well they felt they had been prepared for the rigours of classroom teaching; three, to be able, after analysing the data gathered, to recommend ways of improving the on-going Whānau Rūmaki programme.

text: Teachers' perceptions of the effectiveness of the Whānau Rūmaki teacher education programme, 1990-1992 In 1990 the School of Education, the University of Waikato, initiated a special option within its regular pre-service teacher training programmes, an option designed to prepare teachers to deliver the New Zealand curriculum in bilingual and total immersion schools through the medium of te reo Māori. This alternative programmed, designated Whānau Rūmaki, was experimental and evolutionary. Inevitably particular elements of it had to be varied with each year's operation as its architects attempted 'to get the mix right'. It is difficult, however, when engaged in the day to day running of a programme, to stand back and review with any degree of objectivity its strengths and weaknesses. This study, therefore, in retrospect attempts to do three things: one, to follow the initial Whānau Rūmaki student cohort through its three or four year training cycle noting especially the difficulties confronting individual students some of whom dropped out of the programme; two, to gather information from as many as possible of the graduates who graduated about how well they felt they had been prepared for the rigours of classroom teaching; three, to be able, after analysing the data gathered, to recommend ways of improving the on-going Whānau Rūmaki programme.

year: 1997