title: The DNA Binding Protein Lsr2 from Mycobacterium tuberculosis

authors: Summers, Emma Louise

subjects: DNA binding protein

abstract: Lsr2 is a small, basic DNA binding protein that is highly conserved in mycobacteria and related actinomycetes. Lsr2 is essential for growth in Mycobacterium tuberculosis and previous studies have shown that Lsr2 is involved in down-regulating a wide range of genes involved in cell wall synthesis and metabolic functions. This regulatory function is likely to influence bacterial growth and survival.

This research investigated the biochemistry and 3D structure of Lsr2 from M. tuberculosis. Transmission electron microscopy (TEM) analysis of Lsr2 in complex with DNA revealed a regular fibril-like arrangement of protein coating double-stranded DNA. In addition, it was shown that Lsr2 physically protected DNA from DNase activity. The structure of the C-terminal DNA binding domain of Lsr2, determined by others part way through this research, prompted site directed mutagenesis of residues proposed to interact with DNA. Modification of arginine residues significantly reduced the binding of Lsr2 to DNA and fibril-like structures were not observed using TEM, for arginine mutants.

The first crystal structure of the N-terminal domain of Lsr2 is reported here. Two high resolution structures in monoclinic and hexagonal space groups were solved using X-ray crystallography and ab initio phasing. Proteolytic processing of the N-terminus of Lsr2 was revealed by the structure in P2₁ and this process was recreated using the protease trypsin which resulted in crystal formation in a P3₁21 space group. Both structures show linear chains of dimeric N-terminal Lsr2, as shown by crystallographic symmetry, linked by overlapping anti-parallel β-sheets, revealing a mechanism of protein oligomerisation. Oligomerisation only occurs after the removal of the first three residues M1, A2 and K3. In solution, protein oligomerisation was recreated with trypsin, resulting in the formation of large protein complexes. A change in DNA topology after the addition of trypsin to full-length Lsr2/DNA complexes was observed using TEM. This mechanism is likely to be important to M. tuberculosis under “stress” conditions where proteases are known to be upregulated and where cross-linking and condensation of DNA is critical.

text: The DNA Binding Protein Lsr2 from Mycobacterium tuberculosis Lsr2 is a small, basic DNA binding protein that is highly conserved in mycobacteria and related actinomycetes. Lsr2 is essential for growth in Mycobacterium tuberculosis and previous studies have shown that Lsr2 is involved in down-regulating a wide range of genes involved in cell wall synthesis and metabolic functions. This regulatory function is likely to influence bacterial growth and survival.

This research investigated the biochemistry and 3D structure of Lsr2 from M. tuberculosis. Transmission electron microscopy (TEM) analysis of Lsr2 in complex with DNA revealed a regular fibril-like arrangement of protein coating double-stranded DNA. In addition, it was shown that Lsr2 physically protected DNA from DNase activity. The structure of the C-terminal DNA binding domain of Lsr2, determined by others part way through this research, prompted site directed mutagenesis of residues proposed to interact with DNA. Modification of arginine residues significantly reduced the binding of Lsr2 to DNA and fibril-like structures were not observed using TEM, for arginine mutants.

The first crystal structure of the N-terminal domain of Lsr2 is reported here. Two high resolution structures in monoclinic and hexagonal space groups were solved using X-ray crystallography and ab initio phasing. Proteolytic processing of the N-terminus of Lsr2 was revealed by the structure in P2₁ and this process was recreated using the protease trypsin which resulted in crystal formation in a P3₁21 space group. Both structures show linear chains of dimeric N-terminal Lsr2, as shown by crystallographic symmetry, linked by overlapping anti-parallel β-sheets, revealing a mechanism of protein oligomerisation. Oligomerisation only occurs after the removal of the first three residues M1, A2 and K3. In solution, protein oligomerisation was recreated with trypsin, resulting in the formation of large protein complexes. A change in DNA topology after the addition of trypsin to full-length Lsr2/DNA complexes was observed using TEM. This mechanism is likely to be important to M. tuberculosis under “stress” conditions where proteases are known to be upregulated and where cross-linking and condensation of DNA is critical.

year: 2012

title: Teacher-researcher relationships and collaborations in research.

authors: Cowie, Bronwen

subjects: collaborative research

abstract: To understand the complexity of the classroom in ways that might inform teaching research in needed that explains both why and how something works. Teacher-researcher collaboration is essential if this is to happen. Collaborative work can ensure that research builds on from what teachers know and can do. Researchers working with teachers to address their current concerns are more likely to generate insights into what teachers might do and where they might go next. Collaboration can contribute a warrant for relevance for research findings. At the same teachers deepen and enhance their own practice through engaging in the research process. This paper describes and discusses some approaches to collaboration that have enables researchers and teachers to access a diversity of ideas and expertise to their mutual benefit.

text: Teacher-researcher relationships and collaborations in research. To understand the complexity of the classroom in ways that might inform teaching research in needed that explains both why and how something works. Teacher-researcher collaboration is essential if this is to happen. Collaborative work can ensure that research builds on from what teachers know and can do. Researchers working with teachers to address their current concerns are more likely to generate insights into what teachers might do and where they might go next. Collaboration can contribute a warrant for relevance for research findings. At the same teachers deepen and enhance their own practice through engaging in the research process. This paper describes and discusses some approaches to collaboration that have enables researchers and teachers to access a diversity of ideas and expertise to their mutual benefit.

year: 2010

title: "Everything has shaped you": Contributors to young Māori university students' ability to flourish

authors: Major, Darcie Ngahuia

subjects: Maori

abstract: There are many disparities when considering the health and wellbeing of Māori in Aotearoa today, and the education sector is no different, with Māori less likely to leave school with level 2 certificate or higher compared with non-Māori.  In response to the deficit orientated research on Māori and education, my thesis explores those factors contributing to young Māori tertiary students flourishing. My research includes five students from the University of Waikato who were between the ages of 17-24 years. A multi-method qualitative approach was utilised while engaging a Kaupapa Māori research framework. Participants were found to have a range of interconnected factors which contribute to their ability to flourish both inside and outside of the university.  Of particular importance to my participants was relational connectedness or whanaungatanga within and outside of the tertiary environment, landscapes which offered multiple forms of care, and notions of both collective and individual motivation. Some of the challenges experienced by my participants included a sense of socio-cultural isolation, the minimal inclusion of Māori worldviews and perspectives within the general curriculum, and the personal stress of responding to negative stereotypes of Maori as generally unproductive members of society. Flourishing for my participants is therefore multi-layered, and recognises the interconnected nature of health and wellbeing.

text: "Everything has shaped you": Contributors to young Māori university students' ability to flourish There are many disparities when considering the health and wellbeing of Māori in Aotearoa today, and the education sector is no different, with Māori less likely to leave school with level 2 certificate or higher compared with non-Māori.  In response to the deficit orientated research on Māori and education, my thesis explores those factors contributing to young Māori tertiary students flourishing. My research includes five students from the University of Waikato who were between the ages of 17-24 years. A multi-method qualitative approach was utilised while engaging a Kaupapa Māori research framework. Participants were found to have a range of interconnected factors which contribute to their ability to flourish both inside and outside of the university.  Of particular importance to my participants was relational connectedness or whanaungatanga within and outside of the tertiary environment, landscapes which offered multiple forms of care, and notions of both collective and individual motivation. Some of the challenges experienced by my participants included a sense of socio-cultural isolation, the minimal inclusion of Māori worldviews and perspectives within the general curriculum, and the personal stress of responding to negative stereotypes of Maori as generally unproductive members of society. Flourishing for my participants is therefore multi-layered, and recognises the interconnected nature of health and wellbeing.

year: 2017