182
Chapter 5
Production of biomass by reaction 2 follows the stoichiometry of (1) in Problem 5.2. Note that everything
until AcCoA is the same as in Problem 5.2 except that acetoin (CH
3
CHOH-CO-CH
3
) is formed in reaction 7
by condensation of two pyruvate molecules followed by decarboxylation (no NADH or ATP involved).
Entry into the solvent/acid pathways starts with Acetoacetyl-CoA (CHjCOCILCO-S-CoA) formed by
condensation of Acetyl-CoA (CH
3
CO-S-C
0
A) in reaction
8
. Acetoacetyl-CoA may either decarboxylate to
acetone
(CH
3
COCH
3
)
in
reaction
9
or
be
reduced
(reaction
10)
to
buturyl-CoA
(BuCoA)
(CH
3
CH
2
CH
2
CO-S-C
0
A). Reaction 9 requires ATP since 1 C-mole of BuCoA is formed from butyrate for
every C-mole of Acetoacetyl-CoA converted. BuCoA can be either reduced to butanol in reaction 11 or
hydrolyzed to butyrate in reaction 12 (similar to reaction
6
, and again one ATP is produced per molecule of
CoA, i.e., one-fourth ATP per C-mole of BuCoA). In the decarboxylation of pyruvate to Acetyl-CoA
(reaction 3) in C.
acetobutylicum,
pyrovate-ferredoxin oxido reductase acts as electron receptor
(Fdm
Fd
„d).
There are two ways of reoxidation of
Fd,^:
Either NAD' is reduced to NADH (the feature used in Problem
5.2) or 2H* can be reduced to form H2. This last reaction is included as a separate net reaction: NADH
NAD' + H
2
(reaction 13) to account for the considerable amounts of free H
2
produced, especially at high pH
(the acid forming process). At low pH, less H: and more reduced solvents (e.g., butanol) are produced (the
solvent forming process).
a.
For each reaction Vj to vl3, write up the stoichiometry and include the ATP/NADH produced
or consumed. For all carbon-containing compounds the stoichiometry should be on the basis
of 1 C-mole consumed. Reactions 7,
8
, 9 and 12 do not involve any NADH/NAD
conversion. 0.25 ATP is liberated per C-mole converted in reaction 12, and 0.25 ATP is
consumed per C-mole converted in reaction 9. Reactions 7,
8
, 10, 11 and 13 do not involve
ATP/ADP conversion.
Assume that
rm, = rAcCoA
=
rAcaoAcCoA
=
rBuCoA
= ^nadh = rATP are all zero. Determine the
minimum number of measurements needed to observe the system.
b.
Show that the system is observable if
r„ r„ r„
racn, rHBu, rac, and
rb
are the measured rates.
Calculate the remaining nonzero rates as linear combinations of the measured rates.
c.
If v
8
is set to zero, none of the metabolites that are progenies of AcetoAcCoA are formed. If,
furthermore, v
7
= 0 the whole pathway is the one considered in Problem 5.2. Show that the
expressions for rHAc, rniac, and rc are the same in the degenerate version of Problem 5.2 if
rll:
= 0.
d.
Reardon
et al.
(1987), Fig. 4, gives some results from a batch fermentation carried out at pH =
6
. Between 10 and 15 h after the start of the batch the glucose concentration s and the
concentrations of HAc, HBu, and cells are all fitted well by linear functions of fermentation
time. Virtually no acetoin, lactate, ethanol, acetone, or butanol is produced. CO
2
and H
2
production were not measured. The following concentration changes can be read from the
figure (with at least 5% uncertainty) for the 5-h period:
Glucose
-117mM(=-21.1 g/L)
HBu
76 mM (= 6.69 g/L)
HAc
55 mM (= 3.30 g/L)
Biomass
3.3 g dry weight/L
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