Participation of an nitric oxide synthase in the oxygen dependent
regulation of fermentation in Pichia stipitis.
Ulrich Klinner (1), Magnus Mergler (1), Volkmar Passoth (2)
(1) Institut für Biologie IV, RWTH Aachen, Worringer Weg,
Aachen, 52056, Germany; (2) Swedish University of Agricultural Sciences
(SLU), Dept. of Microbiology, Uppsala Genetic Centre, Box 7025, S-750 07
Uppsala, Sweden (ulrich.klinner@rwth-aachen.de)
During aerobic cultivation of P. stipitis,
alcohol dehydrogenase (ADH) and pyruvate decarboxylase (PDC) activities
are induced after shifting the oxygen tension down from 80 to 20% [1].
However, induction of oxygen dependent systems of most organisms occurs
at an oxygen tension of < 1%, because the Km of these systems
towards oxygen is < 3 microM. Nitric oxide synthases (NOS) are
O2-dependent systems with a higher Km. The NO-radical is well known from mammals as a signalling molecule with the
soluble guanylate cyclase as receptor [2]. The only yeast in which an
NOS activity was described so far is C. tropicalis [3]. We
studied PDC activity of CBS 5776 during cultivations with controlled
aeration and observed activation during aerobic growth (between 94 and
82% oxygen tension) and, more distinctly, after a shift from 80 to 20%.
Sodium nitroprusside, an NO releaser used as a vascular dilating drug in
medicine, had no effect on PDC activity in aerobic cultures.
Surprisingly, it increased PDC activation after the shift up to
fourfold. The formation of the NOS product 14C-citrulline
from 14C-arginine was detected in cell extracts of P.
stipitis. As in mammals, the citrulline formation was inhibited by
the arginine analog L-NAME but not by D-NAME. [1] Passoth V, Zimmermann
M, Klinner U (1996) Appl Biochem Biotechnol 57-8, 201-212 [2] Denninger
JW, Marletta MA (1999) Biochim Biophys Acta-Bioenerg 1411, 334-350 [3]
Wilken M, Huchzermeyer B (1999) Eur J Cell Biol 78, 209-213