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Recombinant human haemoglobin A (rHbA) was produced by a leucine-requiring strain of Hansenula polymorpha which had been transformed with an integration vector containing the Saccharomyces cerevisiae LEU2 gene and cDNAs for the expression of alpha and beta globin each driven by the H. polymorpha MOX promoter. After 40 generations in a chemostat it was found that the integrated vector had become amplified in the host strain. In some cases this led to an increase in LEU2 gene dosage, but a loss of globin expression cassettes. In other cases the globin gene dosage also increased. These changes coincided with an increase in rHbA production in the culture, which was reversed when the dilution rate was increased. Isolates from a chemostat culture producing elevated levels of rHbA were grown in fed-batch fermentations, resulting in higher productivities than when inoculated with the parent strain. The rHbA produced was purified and characterized. Oxygen binding studies and electrospray mass spectrometry showed that the rHbA had been processed and assembled correctly, and behaved as a fully functional co-operative tetramer.

This paper examines the emerging phenomenon of climate emergency declarations. We focus on the case of Victoria Australia and the 30 councils who have declared a climate emergency with a particular focus on three councils. We explore the drivers, meanings, and implications and to what extent the subsequent plans reflect a reframing of local government roles and actions. We find the emergency declaration movement is catalysing councils beyond symbolic declarations potentially opening up space for change and disruption. Of interest in this paper is also the principal and theoretical implications for citizens, local government, and for research that is connected with this emerging trend. We highlight conclusions, ideas, and perspectives that can be drawn from this study of the Australian practice of climate emergency declarations.