E12 | YEAST METABOLISM
catabolism & anabolism
- we do produce ATP, and H2O, Co2, NH3 (energy-poor end products)
from catabolism
- exergonic (release energy
anabolism requires energy (endergonic) to form large, complex molecules
(buildup) from small
glucose {isomerase} fructose
chew gum, has fructose. skips the isomerization step, committed steps na
agad.
Saccharomyces cerevisiae
objectives
- metabolic fates of pyruvate
- krebs cyvce
- aerobic metabolism
-
- metabolic fates of pyruvate (an alphaketo acid)
2,4-DNPH (+ for alde, ket)
+ result: orange/yellow ppt
-: orange to red solution
terminal (outside) lone pair is the first to attack
if acidic, will not attack. because saturated with H+
DNPH detects pyruvate presence.
in basic (pH 8), pyruvate decarboxylase is inhibited. pyruvate is
not consumed.
incubate yeast in yeast, gluc in diff buffers, diff pH condition.
expect to produce pyruvate. at what condition does the
pyruvate change? if pyruvate decarboxylase inhibited, pyruvate
not consumed, ppt produced.
2,4-DNPH positive in blank (w gluc) only if at high temp.
PYRUVATE DECARBOXYLASE ACTIVE AT LOW PH OR
NEUTRAL.
effect of pH
- fermentation favored when acidic. ethanol produced.
o can also TCA cycle
- neutral
o normal condition, TCA cycle (pyruvate consumed)
- basic (high pH)
o pyruvate decarboxylase inhibited
TCA CYCLE
succ fumarate
E: succinate dehydrogenase; FADH2 released
in the expt: want to observe inhibiting effect of maleic, activating effect of
phenol
determined by methylene blue as a redox indicator
reduced to colorless when activated
if inhibited, remains blue.
we look at production of CO2. when it is produced after
consumption of glucpyruvate etc, CO2 gives reducing envt for
methylene blue.
more CO2 production = pyruvate goes into TCA (metaebolism
occurs) = methylene blue becomes colorless
phenol activates. provides mildly acidic envt for solution, favors
TCA. thus, CO2 produced. thus, reducing environment, so met blue
reduced, becomes colorless.
maleic acid is an inhibitor of succinic acid. similar structure-wise,
are analogous and thus compete for succinate dehydrogenase.
o thus, only 2 CO2 formed (TCA does not proceed further)
fructose decolorizes more quickly than glucose. skips isomerization part into
fructose. next, PFK to F-1,6-P. rate of decolorization differs, but the color
should be the same if left for a while.
EFFECT OF ENERGY SRC AND TEMPERATURE
TEMPERATURE 25 vs 37 vs 60
37 deg C optimal. body temp.
at 60 deg, proteins get denatured. no more CO2 production. wala
talaga.
(at 25 deg C, more on reproduction of yeast lang ang favored. some
production of CO2 (some glycolysis), but slowwww)
37 > 25 > 60
ENERGY SOURCE glucose starch sucrose
in terms of rate
glucose > sucrose > starch
sucrose has gluc, fruc. starch is more diff to break down. it needs
high temp, acid hydrolysis, etc, to break glycosidic bond.
glucose, on the other hand, needs no hydrolysis.
sucrose faster than starch. starch is basically made up only of glucose.
sucrose, once hydrolyzed, has both fruc and gluc (fruc is faster), thus,
paglaki ng balloon is bigger in suc compared to starch. easier to
hydrolyze
in the long run, sucrose has biggest balloon. it has both gluc and fruc
units.
EFFECT OF INHIBITORS
sodium fluoride
fluoride ion sequesters Mg2+
formation constant of MgF2 is higher. aagawin yung Mg2+ from
the enzyme. (many enzymes are metalloenzymes)
sodium citrate
- inhibits PFK (phosphofructokinase)
o F-6P[PFK] F-1,6-BP COMMITTED STEP. tuluy-tuloy na sa
glycolysis at this point.
o allosteric regulator.
sodium citrate attaches to PFK; PFK can no longer
phosphorylate F-6P
CITRATE IS A BETTER INHIBITOR? DEBATABLE. NO ACTUAL TREND. JUST BASE
IT ON YOUR DATA.
BIGGER SLOPE, WORSE INHIBITOR.
E1~E4 >> E2,3,5,6
PROBSET
