title: Cellulases from extremely thermophilic anaerobic bacteria: a comparison of several new cellulolytic isolates and the partial purification and characterisation of components of the cellulase complex from one isolate

authors: Sharrock, Keith R.

abstract: Forty-seven thermal pool sites in the central volcanic plateau of the North Island of New Zealand were sampled and enrichment cultures on crystalline cellulose screened for growth and cellulase activity. Eight anaerobic cellulolytic cultures resulted. Cellulolytic isolates were obtained from five of these cultures. All were obligate anaerobic non-sporing rods, staining Gram-negative. Most grew well at 75°C but none grew at 80°C. CMCase activity from some isolates exhibited exceptional thermal stability, with half-lives at 85°C in excess of 10 hours. Isolates from two sources, designated TP8 and TP10, were capable of completely hydrolysing crystalline cellulose and accumulated reducing sugar and avicelase activity in the growth medium. Levels of CMCase and avicelase in our most active culture supernatants were between 10 and 35% of those of Clostridium thermocellum grown on the same medium (but at 60°C). All produced acetic acid and ethanol in varying proportions, with a consequent tendency to decrease the pH of the growth medium. Cellobiose at 0.2%(w/v) repressed production of CMCase activity by most of the isolates by about 50%, but led to a slightly increased CMCase production by two of them.
One of the isolates capable of completely hydrolysing crystalline cellulose (TP8.T6.3.3.1) was subjected to repeated subculture, reselection and reisolation to ensure its purity. The cell-free cellulase complex of this organism was found to produce partial solubilisation of a range of natural cellulosics, and also contained β-glucosidase activity.
Ammonium sulphate precipation and adsorption onto crystalline cellulose were the best of the methods tested for concentrating the low levels of cellulolytic activity in the culture supernatant of TP8.T6.3.3.1. Adsorption onto cellulose was the cheapest alternative and also doubled as a preliminary fractionation and purification step.
Three fractions, separated on the basis of their affinities for crystalline cellulose, were found to interact synergistically in hydrolysing crystalline cellulose when recombined.
 Conventional ion exchange and gel permeation column chromatography served only to subdivide the TP8.T6.3.3.1 cellulase complex into subcomplexes. These were fractionated further using preparative SDS-PAGE and IEF. The cellulose-binding component of the cellulase complex was found to contain an unexpectedly large number of active components (20 being a conservative estimate). Several of these component proteins exhibited clear differences in relative reactivities towards a range of cellulosic substrates. When rerun on SDS-PAGE, almost all produced single protein bands with mobilities indicative of molecular weights covering the range 20,000 to 140,000. Isoelectric points were all in the range pH 4.5-5.0. However, narrow range isoelectric focussing (pH 4.5-5.0) produced further subdivision of these single bands. The bands produced by isoelectric focussing (7-10 per SDS-PAGE band) each bore a share of the endoglucanase activity. We were unable to individually characterize the component activities at this level, and it was not clear whether these multiple components were artefacts of the isoelectric focussing, distinct products of multiple genes, modified products of a relatively small number of genes or microheterogeneity due to ageing of the proteins.

text: Cellulases from extremely thermophilic anaerobic bacteria: a comparison of several new cellulolytic isolates and the partial purification and characterisation of components of the cellulase complex from one isolate Forty-seven thermal pool sites in the central volcanic plateau of the North Island of New Zealand were sampled and enrichment cultures on crystalline cellulose screened for growth and cellulase activity. Eight anaerobic cellulolytic cultures resulted. Cellulolytic isolates were obtained from five of these cultures. All were obligate anaerobic non-sporing rods, staining Gram-negative. Most grew well at 75°C but none grew at 80°C. CMCase activity from some isolates exhibited exceptional thermal stability, with half-lives at 85°C in excess of 10 hours. Isolates from two sources, designated TP8 and TP10, were capable of completely hydrolysing crystalline cellulose and accumulated reducing sugar and avicelase activity in the growth medium. Levels of CMCase and avicelase in our most active culture supernatants were between 10 and 35% of those of Clostridium thermocellum grown on the same medium (but at 60°C). All produced acetic acid and ethanol in varying proportions, with a consequent tendency to decrease the pH of the growth medium. Cellobiose at 0.2%(w/v) repressed production of CMCase activity by most of the isolates by about 50%, but led to a slightly increased CMCase production by two of them.
One of the isolates capable of completely hydrolysing crystalline cellulose (TP8.T6.3.3.1) was subjected to repeated subculture, reselection and reisolation to ensure its purity. The cell-free cellulase complex of this organism was found to produce partial solubilisation of a range of natural cellulosics, and also contained β-glucosidase activity.
Ammonium sulphate precipation and adsorption onto crystalline cellulose were the best of the methods tested for concentrating the low levels of cellulolytic activity in the culture supernatant of TP8.T6.3.3.1. Adsorption onto cellulose was the cheapest alternative and also doubled as a preliminary fractionation and purification step.
Three fractions, separated on the basis of their affinities for crystalline cellulose, were found to interact synergistically in hydrolysing crystalline cellulose when recombined.
 Conventional ion exchange and gel permeation column chromatography served only to subdivide the TP8.T6.3.3.1 cellulase complex into subcomplexes. These were fractionated further using preparative SDS-PAGE and IEF. The cellulose-binding component of the cellulase complex was found to contain an unexpectedly large number of active components (20 being a conservative estimate). Several of these component proteins exhibited clear differences in relative reactivities towards a range of cellulosic substrates. When rerun on SDS-PAGE, almost all produced single protein bands with mobilities indicative of molecular weights covering the range 20,000 to 140,000. Isoelectric points were all in the range pH 4.5-5.0. However, narrow range isoelectric focussing (pH 4.5-5.0) produced further subdivision of these single bands. The bands produced by isoelectric focussing (7-10 per SDS-PAGE band) each bore a share of the endoglucanase activity. We were unable to individually characterize the component activities at this level, and it was not clear whether these multiple components were artefacts of the isoelectric focussing, distinct products of multiple genes, modified products of a relatively small number of genes or microheterogeneity due to ageing of the proteins.

year: 1985

title: Childfree couples' experiences of stereotyping, harassment and pressure

authors: Riley, Theresa Mary

subjects: childfree

abstract: This qualitative study is about New Zealand couples who, by choice, do not have children. Strong social norms exist for couples to have children, and those who express a desire to do otherwise have been disbelieved, pressured, and stereotyped. Womanhood has continued to be associated with motherhood, and a maternal instinct is expected to drive women to have children. The aim of this research was to add to knowledge and awareness of how childfree people have experienced being stereotyped, pressured and harassed for being childfree. For this research, I conjointly interviewed ten heterosexual, childfree couples residing in the city of Hamilton. Participants self-identified as childfree, and ranged in age from 23 to 56 years old. Five of the couples also participated in a focus group. Participants related the ways in which they perceived that the wider social context played a role in the negative responses they experienced. Participants revealed how they felt less socially valued through: an idealization of parenthood, exclusion from work benefits, and an expectation that women should manage both employment and motherhood. Stereotyping was found to still occur, with participants reporting that they were labelled as selfish, immature, and anti-children. Stereotypes of being destined for loneliness in later life, and of their pets being substitutes for children were common. Some evidence was found in participants' comments that there were elements of truth in stereotypes of the childfree. The negative stereotyping appeared to have little, if any, impact on how participants viewed or felt about themselves. Participants reported feeling harassed by other people's disbelief in their choice, and assumptions, that despite what they said, everybody wanted children. The pressures experienced by participants took various forms, such as persistent questioning, and came from various sources, including siblings and acquaintances. Participants' reports of feeling pressured or harassed seem to reflect minor and fleeting feelings, rather than a continuing concern. How pressuring comments were perceived by the recipient was very context-dependent. Participants tolerated and coped with people's negative responses by various methods, such as confronting, ignoring, and avoiding the topic of children with certain people. I recommend that further research be done, and that work is needed to promote both acceptance of the childfree option, and freedom of women's identification from association with childbearing.

text: Childfree couples' experiences of stereotyping, harassment and pressure This qualitative study is about New Zealand couples who, by choice, do not have children. Strong social norms exist for couples to have children, and those who express a desire to do otherwise have been disbelieved, pressured, and stereotyped. Womanhood has continued to be associated with motherhood, and a maternal instinct is expected to drive women to have children. The aim of this research was to add to knowledge and awareness of how childfree people have experienced being stereotyped, pressured and harassed for being childfree. For this research, I conjointly interviewed ten heterosexual, childfree couples residing in the city of Hamilton. Participants self-identified as childfree, and ranged in age from 23 to 56 years old. Five of the couples also participated in a focus group. Participants related the ways in which they perceived that the wider social context played a role in the negative responses they experienced. Participants revealed how they felt less socially valued through: an idealization of parenthood, exclusion from work benefits, and an expectation that women should manage both employment and motherhood. Stereotyping was found to still occur, with participants reporting that they were labelled as selfish, immature, and anti-children. Stereotypes of being destined for loneliness in later life, and of their pets being substitutes for children were common. Some evidence was found in participants' comments that there were elements of truth in stereotypes of the childfree. The negative stereotyping appeared to have little, if any, impact on how participants viewed or felt about themselves. Participants reported feeling harassed by other people's disbelief in their choice, and assumptions, that despite what they said, everybody wanted children. The pressures experienced by participants took various forms, such as persistent questioning, and came from various sources, including siblings and acquaintances. Participants' reports of feeling pressured or harassed seem to reflect minor and fleeting feelings, rather than a continuing concern. How pressuring comments were perceived by the recipient was very context-dependent. Participants tolerated and coped with people's negative responses by various methods, such as confronting, ignoring, and avoiding the topic of children with certain people. I recommend that further research be done, and that work is needed to promote both acceptance of the childfree option, and freedom of women's identification from association with childbearing.

year: 2008

title: Colonial versus ancestral legacy: The assertion of culture and identity through public arts in Kirikiriroa/Hamilton

authors: Kiff, Shirin

subjects: Ethnography

abstract: This thesis is an ethnography of Hamilton. It focuses on the role of public arts as a cultural regeneration tool and their function in shaping the community(s) in Kirikiriroa/Hamilton through building a sense of place and identity. As a qualitative study, I conducted nine semi- structured interviews with public art artists, art advocates and residents to explore the relational context within which the development and transmission of meaning takes place, a process which is central to culture and identity. The primary purpose of the research interviews was to gain insight into the factors that motivate arts organisers and planners to direct valuable limited resources into public art projects and its observable outcomes. The second purpose of the interviews, conducted while walking to public artworks in the city centre with six participants, was to examine the effect of public art on the public.
My methods also include tracking press reviews, monitoring comments and posts on social media and institutional debates involving residents, artists, and public authorities concerning public arts. I also recorded data by taking photographs and recording field notes of my observations during and after the fieldwork process. Data gathering was also enhanced by the large-scale collection of written materials and resources, including public art publications and HCC policy documents. Taken together, these research methods allowed me to investigate whether Hamilton had achieved its goals in its cultural regeneration efforts. In my analysis, I discuss the role of public art and its association with place and consider how artworks may antagonise Māori-Pākehā relationships or otherwise emphasis residents’ sense of place.
Biculturalism and monoculturalism narratives appear to dominate public arts with the purpose of invoking either a colonial legacy or ancestral legacy while the politics of multiculturalism continue to jockey for place. Colonial legacy refers to the glorifying of colonial achievements while ancestral legacy in this context refers to Māori assertion of history, culture and identity. Particularly, in this research, ancestral legacy belonging to local hapū, Ngāti Wairere. Participants’ narratives reveal four overarching themes, 1) the publics’ resistance to the making of colonial places through public art, 2) Ngāti Wairere struggles to assert ancestral legacy through public arts, 3) the need for ethnically diverse artworks to create and maintain ethnically diverse places, and 4) a yearning for a collective identity represented through public arts. Hamilton’s public artworks are variously perceived as a superficial representation of Māori culture, identity and history, a reminder of a colonial binary discourse, the colonised versus the coloniser, an assertion of either a Māori ancestral legacy or a colonial legacy or, minimally, a nod to multiculturalism. Nevertheless, my research shows that participants engagement with these artworks reveal a yearning for a local collective identity that is still a work in progress.

text: Colonial versus ancestral legacy: The assertion of culture and identity through public arts in Kirikiriroa/Hamilton This thesis is an ethnography of Hamilton. It focuses on the role of public arts as a cultural regeneration tool and their function in shaping the community(s) in Kirikiriroa/Hamilton through building a sense of place and identity. As a qualitative study, I conducted nine semi- structured interviews with public art artists, art advocates and residents to explore the relational context within which the development and transmission of meaning takes place, a process which is central to culture and identity. The primary purpose of the research interviews was to gain insight into the factors that motivate arts organisers and planners to direct valuable limited resources into public art projects and its observable outcomes. The second purpose of the interviews, conducted while walking to public artworks in the city centre with six participants, was to examine the effect of public art on the public.
My methods also include tracking press reviews, monitoring comments and posts on social media and institutional debates involving residents, artists, and public authorities concerning public arts. I also recorded data by taking photographs and recording field notes of my observations during and after the fieldwork process. Data gathering was also enhanced by the large-scale collection of written materials and resources, including public art publications and HCC policy documents. Taken together, these research methods allowed me to investigate whether Hamilton had achieved its goals in its cultural regeneration efforts. In my analysis, I discuss the role of public art and its association with place and consider how artworks may antagonise Māori-Pākehā relationships or otherwise emphasis residents’ sense of place.
Biculturalism and monoculturalism narratives appear to dominate public arts with the purpose of invoking either a colonial legacy or ancestral legacy while the politics of multiculturalism continue to jockey for place. Colonial legacy refers to the glorifying of colonial achievements while ancestral legacy in this context refers to Māori assertion of history, culture and identity. Particularly, in this research, ancestral legacy belonging to local hapū, Ngāti Wairere. Participants’ narratives reveal four overarching themes, 1) the publics’ resistance to the making of colonial places through public art, 2) Ngāti Wairere struggles to assert ancestral legacy through public arts, 3) the need for ethnically diverse artworks to create and maintain ethnically diverse places, and 4) a yearning for a collective identity represented through public arts. Hamilton’s public artworks are variously perceived as a superficial representation of Māori culture, identity and history, a reminder of a colonial binary discourse, the colonised versus the coloniser, an assertion of either a Māori ancestral legacy or a colonial legacy or, minimally, a nod to multiculturalism. Nevertheless, my research shows that participants engagement with these artworks reveal a yearning for a local collective identity that is still a work in progress.

year: 2021