title: The Biology and Biochemistry of PhoH2 proteins

authors: Andrews, Emma Sophie Vout

subjects: Mycobacteria

abstract: PhoH2 proteins are found in a diverse range of organisms. To date little is known about these proteins and the role they play in the organisms in which they reside. PhoH2 is a PIN PhoH domain fusion, and these proteins are currently annotated as having unknown function and are described as PhoH like. PhoH domains are thought to be ATPases and all characterised PIN domain proteins are RNases. Most efforts have focussed on determining the role of PIN domain proteins that comprise the toxic component (VapC) of VapBC toxin antitoxin systems, in which the PIN domain is coexpressed as part of an operon with an inhibitor (VapB). In the remaining cases where PIN domain proteins can be found such as PIN PhoH domain fusions, these cases remain unexamined. This thesis describes the biological and biochemical characterisation of the PIN PhoH protein, PhoH2 from Mycobacterium tuberculosis and Mycobacterium smegmatis, along with a preliminary structural characterisation of a thermophilic PhoH2 protein homologue. 

The phoH2 gene from both mycobacterial organisms was found to be expressed as part of a long mRNA transcript. Examination of these transcripts revealed possible alternative 5’ start sites out of frame with the phoH2 gene. For protein overexpression, and ‘normal’ growth and colony formation with conditional overexpression, phoH2 from M. tuberculosis required 152 bp of the 5’ sequence directly upstream of the annotated phoH2 gene (annotated here as phoH2alt). PhoH2 proteins: PhoH2alt MTB, PhoH2alt MSMEG and PhoH2MSMEG show ATP/Mg²⁺ dependent, sequence specific RNA unwinding and cleavage. The sequence (A C) (A/U) (A/U) (G/C) U was deduced as a substrate for PhoH2, and PhoH2alt MTB also demonstrated unwinding and cleavage activity on its upstream ~152 base RNA transcript, suggesting a potential autoregulatory mechanism. Structural analysis of a thermophilic PhoH2 protein homologue has provided preliminary crystallographic data which along with electron microscopy suggest a ring like hexameric PhoH2 oligomer.

text: The Biology and Biochemistry of PhoH2 proteins PhoH2 proteins are found in a diverse range of organisms. To date little is known about these proteins and the role they play in the organisms in which they reside. PhoH2 is a PIN PhoH domain fusion, and these proteins are currently annotated as having unknown function and are described as PhoH like. PhoH domains are thought to be ATPases and all characterised PIN domain proteins are RNases. Most efforts have focussed on determining the role of PIN domain proteins that comprise the toxic component (VapC) of VapBC toxin antitoxin systems, in which the PIN domain is coexpressed as part of an operon with an inhibitor (VapB). In the remaining cases where PIN domain proteins can be found such as PIN PhoH domain fusions, these cases remain unexamined. This thesis describes the biological and biochemical characterisation of the PIN PhoH protein, PhoH2 from Mycobacterium tuberculosis and Mycobacterium smegmatis, along with a preliminary structural characterisation of a thermophilic PhoH2 protein homologue. 

The phoH2 gene from both mycobacterial organisms was found to be expressed as part of a long mRNA transcript. Examination of these transcripts revealed possible alternative 5’ start sites out of frame with the phoH2 gene. For protein overexpression, and ‘normal’ growth and colony formation with conditional overexpression, phoH2 from M. tuberculosis required 152 bp of the 5’ sequence directly upstream of the annotated phoH2 gene (annotated here as phoH2alt). PhoH2 proteins: PhoH2alt MTB, PhoH2alt MSMEG and PhoH2MSMEG show ATP/Mg²⁺ dependent, sequence specific RNA unwinding and cleavage. The sequence (A C) (A/U) (A/U) (G/C) U was deduced as a substrate for PhoH2, and PhoH2alt MTB also demonstrated unwinding and cleavage activity on its upstream ~152 base RNA transcript, suggesting a potential autoregulatory mechanism. Structural analysis of a thermophilic PhoH2 protein homologue has provided preliminary crystallographic data which along with electron microscopy suggest a ring like hexameric PhoH2 oligomer.

year: 2013

title: Urban narrative: Computational linguistic interpretation of large format public participation for urban infrastructure

authors: Dyer, Mark

abstract: Urban Narrative works at the interface between public participation and participatory design to support collaboration processes for urban planning and design. It applies computational linguistics to interpret large format public consultation by identifying shared interests and desired qualities for urban infrastructure services and utilities. As a proof of concept, data was used from the Christchurch public engagement initiative called ‘Share an Idea,’ where public thoughts, ideas, and opinions were expressed about the future redevelopment of Christchurch after the 2011 earthquakes. The data set was analysed to identify shared interests and desired connections between institutional, communal, or personal infrastructures with the physical urban infrastructures in terms of buildings, public places, and utilities. The data has been visualised using chord charts from the D3 JavaScript open source library to illustrate the existence of connections between soft and hard urban infrastructures along with individual contributions or stories. Lastly, the analysis was used to create an infographic design brief that compares and contrasts qualitative information from public consultation with quantitative municipal statistical data on well-being.

text: Urban narrative: Computational linguistic interpretation of large format public participation for urban infrastructure Urban Narrative works at the interface between public participation and participatory design to support collaboration processes for urban planning and design. It applies computational linguistics to interpret large format public consultation by identifying shared interests and desired qualities for urban infrastructure services and utilities. As a proof of concept, data was used from the Christchurch public engagement initiative called ‘Share an Idea,’ where public thoughts, ideas, and opinions were expressed about the future redevelopment of Christchurch after the 2011 earthquakes. The data set was analysed to identify shared interests and desired connections between institutional, communal, or personal infrastructures with the physical urban infrastructures in terms of buildings, public places, and utilities. The data has been visualised using chord charts from the D3 JavaScript open source library to illustrate the existence of connections between soft and hard urban infrastructures along with individual contributions or stories. Lastly, the analysis was used to create an infographic design brief that compares and contrasts qualitative information from public consultation with quantitative municipal statistical data on well-being.

year: 2020

title: A report to iwi on the kaupapa Māori environmental outcomes and indicators kete

authors: Jefferies, Richard

subjects: New Zealand

abstract: Tangata whenua in Aotearoa have been largely excluded from participation in local government  planning since colonisation, but tikanga and Māori values have for the past two decades been acknowledged in resource management and local government legislation, especially the Resource Management Act, 1991 (RMA) and Local Government Act, 2002 (LGA). For example, the RMA has provisions in over 30 sections for councils to give effect to Māori interests.

In practice, however, there is widespread concern that despite these provisions, Māori are largely excluded from local government resource management processes and their values subordinated to those of the wider community, particularly western scientific values.

This report describes research that resulted in a kaupapa Māori outcomes and indicators framework, and associated methods, that can be used by iwi to assess the quality of statutory plans and the environmental performance of councils in their rohe.

text: A report to iwi on the kaupapa Māori environmental outcomes and indicators kete Tangata whenua in Aotearoa have been largely excluded from participation in local government  planning since colonisation, but tikanga and Māori values have for the past two decades been acknowledged in resource management and local government legislation, especially the Resource Management Act, 1991 (RMA) and Local Government Act, 2002 (LGA). For example, the RMA has provisions in over 30 sections for councils to give effect to Māori interests.

In practice, however, there is widespread concern that despite these provisions, Māori are largely excluded from local government resource management processes and their values subordinated to those of the wider community, particularly western scientific values.

This report describes research that resulted in a kaupapa Māori outcomes and indicators framework, and associated methods, that can be used by iwi to assess the quality of statutory plans and the environmental performance of councils in their rohe.

year: 2009-06-30