P-14
Brewing yeast, gene expression and beer quality. Recent advances in molecular biology will probably deliver novel marker genes
to track sugar metabolism, and secondary flavour pathways to improve quality and
beer stability. The sequence and abundance of mRNA in a cell is directly related
to the genome DNA sequence and the cellular response to the environment. This in
turn determines the type and amount of cellular protein that is produced. The
measurement of mRNA levels provides an elegant way to infer the production of
protein. We have demonstrated that maltose permease gene expression can be
estimated using a probe specific for the MALx1 mRNA. The mRNA expression
reflected observed maltose and maltotriose uptake from the wort and remained
high until all maltose and maltotriose was eliminated. Similarly, other genetic
markers could be, in theory, used to determine and predict various aspects of
yeast performance during beer fermentations. For instance, we have used zinc
limitation to identify genes that are able to report on Zn status within the
cells. Zn induces large changes in the expression of many genes and two in
particular. In microarray GeneFilters® results, the highly homologous genes
YOR387c and YGL258w were identified as having approximately 300-fold higher
expression in zinc deficient conditions than under normal conditions. This was
about 40-fold higher than the known zinc-responsive genes ZAP1 and ZRT1. To
establish whether the regulation of YOR387c and YGL258w were suitable for
effective monitoring of zinc-deficient conditions in industrial processes, a DNA
probe was constructed to quantify mRNA expression levels in various conditions
using Northern analysis. The transcript level of YOR387c/YGL285w mRNA was very
high in zinc-deficient conditions, whereas in zinc-replete medium the transcript
level was not detectable. The mRNA level of ZRT1, a gene known to be induced in
response to zinc-deficient conditions was also increased. The basal level of
ZRT1 expression in medium containing zinc was relatively high compared to the
expression of YOR387c/YGL285w, making ZRT1 less useful as a marker gene. To
determine whether the increase in expression of YOR387c/YGL258w was specific for
zinc deficiency and not a general stress response, gene expression was tested
under oxidative stress and carbon starvation conditions. YOR387c/YGL285w and
ZRT1 expression levels were responsive only to the zinc-depleted conditions. The
heat-shock protein, HSP12, was another gene that was induced in zinc depletion
conditions, but the usefulness of this gene as a marker gene for zinc deficiency
is limited since it is highly induced by other conditions.
Peter Rogers is the manager of new technology, a unit within Operations at
Carlton and United Breweries. He has been with the company since 1997. He has a
background in yeast physiology at one end of the science spectrum, the discovery
end, and at the other, the applied end, of method development for large-scale
production of biologicals from agricultural sources. He is a graduate of the
Australian National University. He is the immediate past president of
AusBioTech, the Australian Biotechnology Association.
Ian Dawes (1), Rachel Day (1), David Duan (2), Felicity Roddick (2), Grant
Stanley (3), Paul Chambers (3), Meredith Chandler (3), Vince Higgins (4) Aldo
Lentini (4), and PETER ROGERS (4). (1) School of Biochemistry and Molecular
Genetics, Univ of New South Wales 2052; (2) Dept of Chemical Engineering, RMIT
3000; (3) School of Life Sciences and Technology, VUT 8001; (4) Carlton and
United Breweries, Abbotsford 3067 Australia.