To investigate the transforming potential of hepatitis C virus (HCV), HCV core protein was produced in BALB/3T3 A31-I-1 cells. The cells expressing HCV core gene cooperatively with the v-H-ras gene showed loss of contact inhibition, morphological alterations, and anchorage-independent and serum-independent growth. The cells producing HCV core protein showed enhanced growth against stimulus of growth factor. In addition, antisense oligodeoxynucleotides against mRNA encoding HCV core protein suppressed the growth of HCV core-producing cells. Furthermore, HCV core protein activated mitogen-activated protein kinase and serum response element, which respond to growth stimuli. From these results, we concluded that HCV core protein is involved in the acquisition of cell growth advantage.
Isoamyl acetate is a major determinant of the quality of Japanese sake.
The amount of isoamyl
acetate in the cultures of Hansenula mrakii and
Saccharomyces
cerevisiae Kyokai No. 7, which
is
industrially used in sake fermentation, and the isoamyl
acetate-producing activities of each yeast
strain were compared to investigate biochemical properties of the
producing system of isoamyl acetate
in these yeast strains. S.
cerevisiae could
not produce, or produced an extremely low level of,
isoamyl
acetate when the cells were cultured under aerobic conditions, while
H. mrakii could produce a
large amount of isoamyl acetate cultured at both 15 and 30 °C under
aerobic conditions. Intact
cells of H.
mrakii cultured at 15 and 30 °C
could produce isoamyl acetate from isoamyl alcohol and
acetic acid. Alcohol acetyltransferase activity of H.
mrakii was detected in insoluble fractions, while
isoamyl acetate-synthesizing esterase was detected only in soluble
fractions of the cell extracts.
Isoamyl acetate-hydrolyzing esterase was detected in both soluble
and insoluble fractions. Expression patterns of esterase were examined by native PAGE followed by
activity staining by using
1-naphthyl acetate and Fast Blue B salt. H. mrakii is
likely to have several esterases, and their
expressions were varied depending upon the growth phase and
temperature.
Keywords: Hansenula yeast; isoamyl acetate; esterase; alcohol
acetyltransferase
Isoamyl acetate is synthesized from isoamyl alcohol and acetyl coenzyme A by alcohol acetyltransferase (AATFase) inSaccharomyces cerevisiae and is hydrolyzed by esterases at the same time. We hypothesized that the balance of both enzyme activities was important for optimum production of isoamyl acetate in sake brewing. To test this hypothesis, we constructed yeast strains with different numbers of copies of the AATFase gene (ATF1) and the isoamyl acetate-hydrolyzing esterase gene (IAH1) and used these strains in small-scale sake brewing. Fermentation profiles as well as components of the resulting sake were largely alike; however, the amount of isoamyl acetate in the sake increased with an increasing ratio of AATFase/Iah1p esterase activity. Therefore, we conclude that the balance of these two enzyme activities is important for isoamyl acetate accumulation in sake mash.
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