title: Structural and enzymatic characterisation of nucleoside triphosphate diphosphohydrolases from Trifolium repens and Dolichos biflorus

authors: Cumming, Mathew Hoani

subjects: NTPDase

abstract: Extracellular nucleoside triphosphate diphosphohydrolases (NTPDases) are enzymes that reduce the extracellular nucleotide signal and inactivate the purinogenic signalling pathway. These enzymes, in the presence of a divalent cation sequentially hydrolyses the γ- and β- phosphoanhydride bond from a range of nucleotide di- or triphosphates to the corresponding nucleotide monophosphate.

There is evidence that purinogenic signalling is present in higher plants and it is becoming clear that NTPDases play a role in the early stages of rhizobium infection during nodulation in legumes. To further our understanding of NTPDases, this study investigated the biochemical and structural characteristics of two legume NTPDases believed to be involved in nodulation. The first, 7WC, was isolated from the roots of white clover and the second, DbLnP, from the roots of Dolichos biflorus. DbLnP has been characterised as a carbohydrate binding NTPDase that is directly associated with the perception of rhizobia. Using X-ray crystallography a number of crystal structures of 7WC and DbLnP were determined at resolutions between 1.9 Å and 2.9 Å. For 7WC, structures were determined for an apo- form, an AMP-bound and also bound with the nonhydrolysable ATP analogue AMPPNP. For DbLnP structures were solved with phosphate and Mn2+ bound and another with AMPPNP and Mn2+ bound. Kinetic analysis of a range of substrates together with the analysis of the binding modes of 7WC and DbLnP explained substrate preference for each of the NTPDases.
These analyses showed that NTPDases can adopt two conformations depending on substrate and co-factor  binding.    The  central  hinge  region  creates  a  ‘butterfly’   motion of the domains that reduces the width of the active site cleft. This phenomenon has been previously hypothesised but has not been observed for NTPDases.
￼￼￼￼￼￼A model of catalysis is  proposed  whereby  the  ‘open’  form  first  binds  substrate  in   an inactive orientation. Binding of the metal ion induces a conformational change that brings the domains together and allows movement of the phosphate tail deeper into the active site cleft – a reorganisation that is required for catalysis. Hydrolysis occurs via nucleophilic attack on the terminal phosphate. Finally, release   of   the   metal   ion   allows   the   ‘open’   conformation   to   be   restored   for   subsequent catalysis to occur.

text: Structural and enzymatic characterisation of nucleoside triphosphate diphosphohydrolases from Trifolium repens and Dolichos biflorus Extracellular nucleoside triphosphate diphosphohydrolases (NTPDases) are enzymes that reduce the extracellular nucleotide signal and inactivate the purinogenic signalling pathway. These enzymes, in the presence of a divalent cation sequentially hydrolyses the γ- and β- phosphoanhydride bond from a range of nucleotide di- or triphosphates to the corresponding nucleotide monophosphate.

There is evidence that purinogenic signalling is present in higher plants and it is becoming clear that NTPDases play a role in the early stages of rhizobium infection during nodulation in legumes. To further our understanding of NTPDases, this study investigated the biochemical and structural characteristics of two legume NTPDases believed to be involved in nodulation. The first, 7WC, was isolated from the roots of white clover and the second, DbLnP, from the roots of Dolichos biflorus. DbLnP has been characterised as a carbohydrate binding NTPDase that is directly associated with the perception of rhizobia. Using X-ray crystallography a number of crystal structures of 7WC and DbLnP were determined at resolutions between 1.9 Å and 2.9 Å. For 7WC, structures were determined for an apo- form, an AMP-bound and also bound with the nonhydrolysable ATP analogue AMPPNP. For DbLnP structures were solved with phosphate and Mn2+ bound and another with AMPPNP and Mn2+ bound. Kinetic analysis of a range of substrates together with the analysis of the binding modes of 7WC and DbLnP explained substrate preference for each of the NTPDases.
These analyses showed that NTPDases can adopt two conformations depending on substrate and co-factor  binding.    The  central  hinge  region  creates  a  ‘butterfly’   motion of the domains that reduces the width of the active site cleft. This phenomenon has been previously hypothesised but has not been observed for NTPDases.
￼￼￼￼￼￼A model of catalysis is  proposed  whereby  the  ‘open’  form  first  binds  substrate  in   an inactive orientation. Binding of the metal ion induces a conformational change that brings the domains together and allows movement of the phosphate tail deeper into the active site cleft – a reorganisation that is required for catalysis. Hydrolysis occurs via nucleophilic attack on the terminal phosphate. Finally, release   of   the   metal   ion   allows   the   ‘open’   conformation   to   be   restored   for   subsequent catalysis to occur.

year: 2012

title: ASEAN and the non-traditional regional security agenda

authors: Rolls, Mark G.

subjects: ASEAN

abstract: Without a doubt, the whole area of non-traditional security – whether it be threats, issues or challenges (the three are more or less used interchangeably in ASEAN statements and documents on the subject) – has come to occupy a prominent place on the regional security agenda in recent years.1 This is very much in keeping with the ‘widening’, or ‘broadening’, of the security agenda which has occurred internationally in both the academic literature and, crucially, in the policy sphere.

text: ASEAN and the non-traditional regional security agenda Without a doubt, the whole area of non-traditional security – whether it be threats, issues or challenges (the three are more or less used interchangeably in ASEAN statements and documents on the subject) – has come to occupy a prominent place on the regional security agenda in recent years.1 This is very much in keeping with the ‘widening’, or ‘broadening’, of the security agenda which has occurred internationally in both the academic literature and, crucially, in the policy sphere.

year: 2010

title: Safety hubs and partnership policing: Coordination, collaboration and partnership working in Auckland city centre

authors: Tompson, Lisa

subjects: Community Safety

abstract: This report presents key insights from an exploratory study of the Safety Hub initiative that first appeared in the run up to the 2023 Women’s World Cup in Auckland. It highlights the potential and the pitfalls of partnership-based approaches to community safety in Aotearoa New Zealand. The study was based on a series of in-person focus group discussions and semi-structured interviews with key partners and stakeholders including Community Patrols New Zealand (CPNZ) volunteers, NZ Police (Auckland city Beat officers and Liaison staff), Auckland council staff and Ngā Watene Māori (Māori Wardens). Data collection for this study took place over a four-month period from November 2024 through to February 2025, structured around the availability of participants. The study employed a thematic analysis methodology to systematically identify and organise key themes and issues within the qualitative data gathered from interviews and focus groups.
In summary, the operational reality of the Safety Hub was rather less structured and more ‘organic’ than some participants had hoped, but it still delivered meaningful outcomes. The Safety Hub made its own valuable contribution to community safety while the difficulties it faced helped identify the need for a wider, linked-up ecosystem of community safety in Auckland city. The Hub has proved itself to be a viable concept, that diverse partners can work together in a shared space, and that the presence of volunteers does offer reassurance and can make people feel safer. However, those involved in the Hub also identified a range of challenges that need to be overcome if the Hub is to be more than just a base for increasing the visibility and presence of capable guardians.

text: Safety hubs and partnership policing: Coordination, collaboration and partnership working in Auckland city centre This report presents key insights from an exploratory study of the Safety Hub initiative that first appeared in the run up to the 2023 Women’s World Cup in Auckland. It highlights the potential and the pitfalls of partnership-based approaches to community safety in Aotearoa New Zealand. The study was based on a series of in-person focus group discussions and semi-structured interviews with key partners and stakeholders including Community Patrols New Zealand (CPNZ) volunteers, NZ Police (Auckland city Beat officers and Liaison staff), Auckland council staff and Ngā Watene Māori (Māori Wardens). Data collection for this study took place over a four-month period from November 2024 through to February 2025, structured around the availability of participants. The study employed a thematic analysis methodology to systematically identify and organise key themes and issues within the qualitative data gathered from interviews and focus groups.
In summary, the operational reality of the Safety Hub was rather less structured and more ‘organic’ than some participants had hoped, but it still delivered meaningful outcomes. The Safety Hub made its own valuable contribution to community safety while the difficulties it faced helped identify the need for a wider, linked-up ecosystem of community safety in Auckland city. The Hub has proved itself to be a viable concept, that diverse partners can work together in a shared space, and that the presence of volunteers does offer reassurance and can make people feel safer. However, those involved in the Hub also identified a range of challenges that need to be overcome if the Hub is to be more than just a base for increasing the visibility and presence of capable guardians.

year: 2025