title: DNA repair in Antarctic bacteria - characterisation of two nuclease proteins from Antarctic dry valley metagenomes

authors: Stelzer, Ronja

subjects: Antarctica

abstract: DNA repair processes are crucial for the growth and survival of all organisms. This is especially true for organisms, that inhabit extreme habitats, like the Antarctic Dry Valleys. The Dry Valleys are one of the coldest and driest desert environments on Earth. The conditions in the Dry Valleys, including high levels of UV radiation, low temperatures, multiple daily freeze-thaw cycles, desiccation, and low nutrient levels, are highly damaging to the genomic DNA of the organisms inhabiting them. The Dry Valleys are dominated by microorganisms, which have a range of adaptations to survive in this extreme environment. Previous studies focus largely on the metabolic adaptation of Dry Valley inhabitants. In this thesis, the DNA repair machinery of bacteria in the Dry Valleys was examined to determine how organisms survive the DNA-damaging conditions.
Our understanding of DNA repair and replication in bacteria pathways is largely based on studies of isolated organisms and pathogens, which have been extensively studied. Methods such as metagenomic sequencing allow us to find novel proteins from currently unculturable organisms. In a search for novel DNA repair proteins in bacterial metagenomes from the Dry Valleys, two unique nuclease proteins were chosen for structural and biochemical characterisation. One of these nuclease proteins belongs to the currently uncharacterised protein domain UPF0102. This protein is distantly related to the archaeal Holliday junction resolvases and is a Type II restriction endonuclease type protein. The activity of DV-Hjc and the UPF0102 domain was characterised in vitro using biochemical assays and in vivo in E. coli knockout cells. Biochemical characterisation of the protein showed that the protein has strong binding affinity with double-stranded DNA substrates and Holliday-junction mimicking substrates. The structure of the protein was determined via X-ray crystallography at a resolution of 0.9 Å and structural features were related to its activity, revealing an extensive DNA binding surface and an active site similar to that of archaeal Holliday junction resolvases. The second nuclease characterised in this thesis is part of a unique nuclease-ligase fusion protein and belongs to the nuclease group of MBL-β-CASP proteins. The biochemical characterisation of the protein showed its ability to bind and cleave DNA, specific activity was observed with abasic site substrates.

text: DNA repair in Antarctic bacteria - characterisation of two nuclease proteins from Antarctic dry valley metagenomes DNA repair processes are crucial for the growth and survival of all organisms. This is especially true for organisms, that inhabit extreme habitats, like the Antarctic Dry Valleys. The Dry Valleys are one of the coldest and driest desert environments on Earth. The conditions in the Dry Valleys, including high levels of UV radiation, low temperatures, multiple daily freeze-thaw cycles, desiccation, and low nutrient levels, are highly damaging to the genomic DNA of the organisms inhabiting them. The Dry Valleys are dominated by microorganisms, which have a range of adaptations to survive in this extreme environment. Previous studies focus largely on the metabolic adaptation of Dry Valley inhabitants. In this thesis, the DNA repair machinery of bacteria in the Dry Valleys was examined to determine how organisms survive the DNA-damaging conditions.
Our understanding of DNA repair and replication in bacteria pathways is largely based on studies of isolated organisms and pathogens, which have been extensively studied. Methods such as metagenomic sequencing allow us to find novel proteins from currently unculturable organisms. In a search for novel DNA repair proteins in bacterial metagenomes from the Dry Valleys, two unique nuclease proteins were chosen for structural and biochemical characterisation. One of these nuclease proteins belongs to the currently uncharacterised protein domain UPF0102. This protein is distantly related to the archaeal Holliday junction resolvases and is a Type II restriction endonuclease type protein. The activity of DV-Hjc and the UPF0102 domain was characterised in vitro using biochemical assays and in vivo in E. coli knockout cells. Biochemical characterisation of the protein showed that the protein has strong binding affinity with double-stranded DNA substrates and Holliday-junction mimicking substrates. The structure of the protein was determined via X-ray crystallography at a resolution of 0.9 Å and structural features were related to its activity, revealing an extensive DNA binding surface and an active site similar to that of archaeal Holliday junction resolvases. The second nuclease characterised in this thesis is part of a unique nuclease-ligase fusion protein and belongs to the nuclease group of MBL-β-CASP proteins. The biochemical characterisation of the protein showed its ability to bind and cleave DNA, specific activity was observed with abasic site substrates.

year: 2024-12-09

title: The cold chain in New Zealand - A review

authors: Carson, James K.

subjects: Science & Technology

abstract: Approximately 45 % of all exports from New Zealand are food products or by-products, and an estimated 60 % of these are exported in the refrigerated state. The leading food sectors in terms of volume and income are in order: the dairy industry, the red meat industry, the horticultural industry and the seafood industry. Due to its geographic isolation, New Zealand food exports form part of some of the longest cold chains in the world. Responsibility for temperature integrity in the cold chain is placed on the processors and exporters and is overseen by the Ministry for Primary Industry of the Government of New Zealand. Currently, the major research theme related to the cold-chain in New Zealand is focussed around optimising chilling and freezing processes for shelf-life extension. Improving energy efficiency in refrigeration processes is also a significant research theme.

text: The cold chain in New Zealand - A review Approximately 45 % of all exports from New Zealand are food products or by-products, and an estimated 60 % of these are exported in the refrigerated state. The leading food sectors in terms of volume and income are in order: the dairy industry, the red meat industry, the horticultural industry and the seafood industry. Due to its geographic isolation, New Zealand food exports form part of some of the longest cold chains in the world. Responsibility for temperature integrity in the cold chain is placed on the processors and exporters and is overseen by the Ministry for Primary Industry of the Government of New Zealand. Currently, the major research theme related to the cold-chain in New Zealand is focussed around optimising chilling and freezing processes for shelf-life extension. Improving energy efficiency in refrigeration processes is also a significant research theme.

year: 2018

title: Indigenous perspectives and gene editing in Aotearoa New Zealand

authors: Hudson, Maui

subjects: Science & Technology

abstract: Gene editing is arguably the most significant recent addition to the modern biotechnology toolbox, bringing both profoundly challenging and enabling opportunities. From a technical point of view the specificity and relative simplicity of these new tools has broadened the potential applications. However, from an ethical point of view it has re-ignited the debates generated by earlier forms of genetic modification. In New Zealand gene editing is currently considered genetic modification and is subject to approval processes under the Environmental Protection Authority (EPA). This process requires decision makers to take into account Māori perspectives. This article outlines previously articulated Māori perspectives on genetic modification and considers the continuing influence of those cultural and ethical arguments within the new context of gene editing. It also explores the range of ways cultural values might be used to analyse the risks and benefits of gene editing in the Aotearoa New Zealand context. Methods used to obtain these perspectives consisted of (a) review of relevant literature regarding lessons learned from the responses of Maori to genetic modification, (b) interviews of selected ‘key Maori informants’ and (c) surveys of self-selected individuals from groups with interests in either genetics or environmental management. The outcomes of this pilot study identified that while Māori informants were not categorically opposed to new and emerging gene editing technologies a priori, they suggest a dynamic approach to regulation is required where specific uses or types of uses are approved on a case by case basis. This study demonstrates how the cultural cues that Māori referenced in the genetic modification debate continue to be relevant in the context of gene editing but that further work is required to characterize the strength of various positions across the broader community.

text: Indigenous perspectives and gene editing in Aotearoa New Zealand Gene editing is arguably the most significant recent addition to the modern biotechnology toolbox, bringing both profoundly challenging and enabling opportunities. From a technical point of view the specificity and relative simplicity of these new tools has broadened the potential applications. However, from an ethical point of view it has re-ignited the debates generated by earlier forms of genetic modification. In New Zealand gene editing is currently considered genetic modification and is subject to approval processes under the Environmental Protection Authority (EPA). This process requires decision makers to take into account Māori perspectives. This article outlines previously articulated Māori perspectives on genetic modification and considers the continuing influence of those cultural and ethical arguments within the new context of gene editing. It also explores the range of ways cultural values might be used to analyse the risks and benefits of gene editing in the Aotearoa New Zealand context. Methods used to obtain these perspectives consisted of (a) review of relevant literature regarding lessons learned from the responses of Maori to genetic modification, (b) interviews of selected ‘key Maori informants’ and (c) surveys of self-selected individuals from groups with interests in either genetics or environmental management. The outcomes of this pilot study identified that while Māori informants were not categorically opposed to new and emerging gene editing technologies a priori, they suggest a dynamic approach to regulation is required where specific uses or types of uses are approved on a case by case basis. This study demonstrates how the cultural cues that Māori referenced in the genetic modification debate continue to be relevant in the context of gene editing but that further work is required to characterize the strength of various positions across the broader community.

year: 2019