Carbon Metabolism Flashcards
Calvin Cycle
also known as Photosynthetic Carbon Reduction Cycle (Reductive Pentose Phosphate Cycle)
Products: PGA, NADP+, ADP
14CO2
distinguish between photosynthetic CO2 and respired CO2 using 4CO2 rationale: expose plant for varying times and identify labeled compounds short time ->early cpds. long time -> late cpds. paper chromatography autoradiography algae - chlorella apparatus
14CO2 experiment
Take a tank of algae, bubble them with CO2, mixture goes to plastic tubing which is illuminated, 14CO2 injected, at the bottom of the tubing, they are dropped in the boiling methanol solution -> autoradiography -> paper chromatography – detect which ones are labeled
—–> labeled compound: PGA (also found in chloroplast)
14CO2 results
after 5 sec PGA (phosphoglyceric acid) is labeled: 3C compound only its carboxyl carbon labeled after 30-90 sec hexose phosphates labeled PGA -- all of its carbons labeled ->What does this suggest?
14CO2 hypothesis:
CO2 + 2C —–light——> PGA
(Assume PGA is the first compound made)
Graph:
1) steady state of 14CO2
2) Turn off the light and see what accumulates
- rxn. above should stop
- What accumulates instead?
>PGA still accumulates (increased)
RUBP disappearing (decreased)
Unexpected result:
CO2 + acceptor doesn’t require light
could RuBP be the acceptor?
rxn doesn’t require light since PGA is accumulating
What is CO2 acceptor? (Old + New hypothesis)
Original hypothesis: two carbon compound?
CO2 + 2 C ---light---> PG
Experiment: steady state with 14CO2
turn lights OFF
PGA accumulates RUBP disappearing
unexpected result
CO2 + acceptor doesn't require light
could RuBP be the acceptor?
RuBP is 5 carbons
New Hypothesis:
CO2 + RuBP —–light—-> 2PGA
(RuBP = ribulose -1, 5 - bisphosphate)
Confirming the acceptor (New Hypothesis)
CO2 + RuBP -----light----> 2PGA
data: steady state with 14CO2
remove CO2; keep lights ON
RuBP increases
PGA disappears
Why? because it is a cycleCalvin Cycle
CO2 + RuBP ——> 2PGA
(1C) (5C) (two 3C = 6C)
3 CO2 + 3 RuBP ————> 6 PGA
(3C) (15 C) (18C)
|
3 RuBP net gain triose-P
(15C) (3C)
used again
Calvin Cycle Process
RuBP and CO2 react in a carboxylation reaction and produce PGA
1 atp and 1 nadph per each PGA –> reduction reaction (ATP + NADPH = NADP+ +ADP) then occurs – > glyceraldehyde3phosphate -> remaining carbon is regenerated
cycle occurs in the stroma inside the cholorplast
triose-P exported (PGA and PGAL) via the
(phosphate - phosphate ester transporter)
2 NADPH : 3 ATP : 1 CO2
Calvin Cycle efficiency
A net gain of 1 hexose requires:
6CO2 + 12 NADPH + 18 ATP
The process is ~90% energy efficient at converting chemical energy into sugars
Autocatalytic
Calvin cycle can be autocatalytic ( if one of the reaction products is also a reactant and therefore a catalyst in the same or a coupled reaction)
Autocatalytic –> 15 Rubp + 15co2 -> 30 PGA -> 30 triose-P -> make 18 RuBP and react again with 15 CO2 and so on – buildup of RuBP OR sugar and starch output
Starch: storage product
made in chlpts
sucrose: storage product
made in cytoplasm from PGA
carbon translocation
Activation of enzymes
Light activation of enzymes
light –> enzymes active
dark –> enzymes inactive
light is not absorbed by enzymes
regulation is indirect
P. E. T. involved
RuBP Carboxylase / Oxygenase (Rubisco)
abundant protein Molecular Weight: 500,000 8 large subunits, 8 small subunits - large subunit chlpts genome catalytic activity - small subunit nuclear genome regulatory activity
Rubisco is the common link between the two pathways (calvin cycle and photorespiration)
activation of RuBP
activity/activation depends on light
enzyme + CO2 (inactive)
|
enzyme--CO2 (inactive)
|
enzyme--CO2--Mg+2 (ACTIVE)
-> maximum activity requires Rubisco activase
readily reversible
depends on [Mg+2] and [CO2]
light dependent activation of Rubisco
Proton (H+) accumulation inside the lumen, Mg2+ kicked out to maintain neutrality
—- > Mg2+ activates Rubisco enzyme
—-> Carbon dioxide is needed to activate rubisco
stromal [Mg+2] increases
stromal pH increases to optimum for Rubisco
lumen pH = 5.0
stroma pH = 8.0
Rubisco activase function
1) Carboxylation
RuBP + CO2-------> 2 PGA
(5 C) (1 C) (6 C)
slow: 3 CO2 / sec
large amount of Rubisco
2) Oxygenation
RuBP + O2 —> PGA + Phosphoglycolate
(5 C) (3 C) (2 C)
O2, CO2 compete for Rubisco active site
[O2] > [CO2] atmospheric concentration
KM (O2) > KM (CO2)
KM: measure of affinity of enzyme for substrate
very slow 1/3 O2 / sec
difficult to measure oxygenase reaction
Photorespiration
(also known as the oxidative photosynthetic carbon cycle, or C2 photosynthesis) refers to a process in plant metabolism where the enzyme RuBisCO oxygenates RuBP, causing some of the energy produced by photosynthesis to be wasted.
Photorespiration (PR)
O2 inhibits photosynthesis – 45% inhibition
lower [O2] – plants grow better
photosynthesis increases
PR - light dependent consumption of
O2 and release of CO2
different from mito respiration
oxygenase is first step in PR stimulated by high [O2] low [CO2] high T : CO2 comes out of water faster than O2 – high O2, low CO2 ratio high light
O2 is consumed
CO2 is released
energy is consumed
eliminating PR
major drain on productivity
2% O2 ——> net photosynthesis 45%
greater than in 21% O2
eliminate PR —-> increase plant productivity
Functions of Photorespiration
1) glycine / serine production (makes amino acids)
2) recycle carbon
oxygenase is unavoidable
not eliminated by evolution
mutants without PR enzymes
die in air
live in high CO2
3) dissipates excess energy under high light and low [CO2]
way to burn off extra energy
Modifying/eliminating PR
a. chemical inhibition:
1970's and 1980's
inhibit enzymes prior to CO2 release
- does it work? NO
1980's
selectively inhibit oxygenase (first step,
prevents formation of enzymes)
can it be done? NO
b. genetic modification – genetically modify Rubisco?
Arabidopsis mutants
mutagenize plant
mutagenize mutants
c. in vitro mutagenesis
clone large subunit
mutagenize; transcribe; translate
look for reduced oxygenase
Genetic modification on PR
- Mutagenize plant to knock out activity of a
photorespiration enzyme
PR enzyme deficient —> plant dies in air - mutagenize mutants
look for plants that live/thrive in air
hope for reduced oxygenase (Rubisco genetically modified)
survivors are revertantsrequires large subunit mutation (chlpts) many genomes/chlpts; several chlpts per cell little chance of multiple mutations
Can reduced oxygenase activity be achieved?
Can the active site of Rubisco be modified to exclude O2 but not CO2?
No, not yet
Photorespiration facts
1) under normal conditions in the field, photorespiration is
not responsible for a significant increase in plant productivity (plant productivity is increased when PR is eliminated)
2) in photorespiration, O2 is consumed and CO2 is released
3) photorespiration occurs within the chloroplast, peroxisome, and mitochondria
4) the first products of the RuBP carboxylation reaction are 2 PGA
5) Products of the RuBP oxygenation reaction are 1 PGA and phosphoglycolate
6) Photorespiration is increased under conditions of high O2, low CO2, high Temp, and high light conditions
