Download Student Questions and Answers October 22, 2002

Student Questions and Answers October 22, 2002
Q 1. What determines if energy is passed by ATP or by GTP? (citric acid cycle)
Answer: FK: To give a simple answer: Of course it´s a matter of specificity of the
respective enzymes, which will either recognise GDP/GTP or ADP at their nucleotide
binding sites. The above question reflects a typical (and unfortunately ongoing)
weakness of textbooks, which give a completely misleading information based on
knowledge of the sixties! Actually most organisms (bacteria, plants, lower animals)
express (only) an ADP-specific succinyl CoA synthetase (SCS), so the “normal” energy
currency is properly used. Only in higher animals two isoforms are expressed, (both
localised in mitochondria), which are specific for GTP and ADP, respectively, and serve
two independent metabolic roles. As in all other organisms, ADP is phosphorylated by
the isozyme which is part of the TCA-cycle, which is energetically possible, since in
mitochondria the ratio of ATP/ADP typically is around 1. On the other hand, GTP/GDP
typically is ~ 100, so you would not expect phosphorylation of GDP to any significant
extent. Indeed, this isoform in vivo catalyses the reverse reaction to replenish succinyl
CoA (from succinate and GTP) when large amounts of it are needed for ketone body
catabolism! (Ketone bodies, i.e. acetoacetate and β-hydroxybutyrate are taken up from
the circulation, but have to be transformed to thioesters to become substrates for the βoxidation system.) Additionally, this GDP-isoform is also involved in porphyrin
synthesis. So it was just bad luck that the first SCS which was characterised was this
isoform, which gave rise to the myth that substrate-level phosphorylation oddly
produces GTP rather than ATP!
Q 2. Why are there 2 currencies of energy, ATP and GTP?
Answer: FK: If each of the 4 ribonucleoside triphosphates is specialised to certain fields
of cellular tasks (which more or less is the case), there is more freedom to independently
control and regulate these activities. (In some sense most triphosphate “energy”
somehow goes back to ATP, which by “trans phosphorylation, catalysed by nucleoside
diphosphokinase, fills up the other triphosphate pools, if necessary.) Unfortunately the
immediate base of this question goes back to some textbook superficiality (see Q 1).
Q 3. When mitochondria multiply, they behave like bacteria. Do they also use ftsz to part the
cells?
Answer: FK: This clearly is a cell biology question! In fact, FtsZ is found in
chloroplasts, but not in mitochondria, where, as was recently shown, it´s function in
septation obviously is taken over by dynamin.
Q 4. In one picture there is a Mg2+ attached to ATP. Is it always attached to ADP and what is
its function?
Answer: FK: The (cluster of) negative charges in ATP and related phosphoryl
compounds provide moderately strong binding of divalent kations. Since intracellular
concentrations of Mg2+ are about 5mM, this complex essentially represents ATP in vivo.
The positive charges of the metal to some degree relieve the electric repulsion stress in
ATP and thus stabilise it (∆G°´ is about 5kJ less negative in the presence of Mg). Most
likely there is no other “function” of this binding.
Q 5. Is there an alternative pathway to glycolysis that can be used by organsims if one or
more enzymes of glycolysis fail?
Answer: FK:. Glycolysis was among the earliest pathways to evolve, all organisms, but
a few bacteria (Rhizobium, Pseudomonas, Azotobacter, Agrobacterium, …), exhibit
glycolysis. These bacteria lack PFK (phosphofructo kinase) and rely on an alternative
pathway called Entner-Doudoroff Pw (GlucoseªGlucose-6-Pª6-P-glucono-δ-lactoneª
6-P-gluconateª2-keto-3-deoxy-6-P-gluconateªpyruvate + glyceraldehyde-3-P (which
is converted to another pyruvate along the “ordinary” route). In total you get 1ATP, 1
NADH, and 1 NADPH per glucose, i.e. it is less efficiently than glucose.
Q 6. Why use ATP as general energy currency if e.g. acetyl-P ª acetate + Pi sets free more
free energy?
Answer: FK: .Ribonucleotides are very old molecules and so nuleotide binding sites on
different proteins are well established, so you can see it as an example for nature´s
economy to use them also as enegy transfer molecules. Developing acetyl phosphatebinding motives would have been an extra task, and not an easy one given the
significantly smaller size of this high energy phosphate compound and the more
pronounced instability of acetyl phosphate. So it is only a by-product of metabolism in
various bacteria, formed from D-xylulose-5-phosphate (ª glyceraldehyde-3-phosphate
+ acetyl phosphate). In a reaction catalysed by phosphate acetyltransferase it transfers
the acetyl group to coenzyme A, giving rise to acetylCoA-formation. Another enzyme
found in many bacteria capable of growing on acetate as sole carbon source is acetate
kinase (acetate + ATP D acetyl phosphate + ADP), again finally acetalCoA is formed.
An additional role in these organisms is in transduction of certain signals (via target
protein acetylation).
Q 7. Is NAD+/NADH used anywhere else but as an e--carrier?
Answer: FK: Only very recently it was established that NAD(P) also serve as precursors
of intracellular signal molecules in 2 different ways: a) by ADP-ribosylation of acceptor
proteins (covalent linkage), and b) via cyclisation (ADP-ribosyl cyclaase acting both on
NAD and NADP; the products cADPR and cADPRP induce Ca-mobilisation from the
ER.
Q 8. Are there other e--carriers than NAD(P)H or FAD(H2)? If yes, which, how do they look
like ? (sorry, can´t read last word)?
Answer: FK: In principle every molecule taking part in redox
reactions is an e--carrier. Obviously you were thinking of
cofactors and prosthetic groups. For sure you already know
several of them, like ubiquinone, plastoquinone, cytochromes,
etc.
Q 9. Why is this hydride ion released? When I consider
electronegativity, C should get the electrons, since it´s
electronegativity is higher.
Answer: Oxidation (i.e. removal of both electrons) of this carbon atom allows
delocalisation of 6 π-electrons through the entire ring, which makes the product much
more stable. However, the way how this is achieved is not as obvious as the outcome,
given the very high reaction rates observed with some NAD-dependent dehydrogenases.
3 types of mechanisms have been proposed for this hydrogen transfer:
a. Direct hydride transfer
b. Free radical mechanisms (transfer of electrons and protons in separate steps)
c. Formation of covalent intermediates between coenzyme and substrate, followed
by electron transfer through these covalent bonds, and transfer of the hydrogen as
a H+
a) and b) are less likely, since they involve high-energy steps and thus should be too
slow for the observed rates. The below presented mechanism (suggested by Hamilton,
1971) therefore most likely best describes what happens.
Q 10. Is it possible to add more than the 3 P groups, e.g. in ATP (forming a tetraphosphate
and so on? If it is possible, why is ATP so dominant in cells? If it is not possible, why?
Answer: FK: Of course; energetically any further P-group would be roughly equivalent
to the β- and γ-P, but this would represent rather an intermediate between rapidly
available energy currency and energy storage molecule (many bacteria in fact use
polyphosphates to store energy). However, for a transient energy transfer molecule this
would only mean an extra load with no increase in efficiency (since the majority of
reactions requires only the tranfer ofa single P-group) and which would make regulation
significantly more complicated.
Q 11. Where does glycolysis take place?
Answer: (You really should already know this! Answer also given in the lecture.) In the
cytosol (anaerobic part). There are rare exceptions: in trypanosomes (unicellular
eukaryotic parasites) the first 7 enzymes of glycolysis are found in unique peroxisomelike organelles, called glycosomes.
Q 12. Where do the e--carriers come from? Where are they located?
Answer: Will be covered in detail, but again you should already be familiar with the
answers.
Q 13. Why isn´t the e- from the first e--donator transferred directly to the final e--acceptor?
Could this occur in the cell?
Answer: FK: (Since I didn´t attend the respective seminar, I am not sure whether I
interpret your question the way it was meant.) a) Nature´s general strategy (and
obviously the only way) to conserve at least a significant fraction of the free energy of
vastly exergonic processes: the direct interaction of the over-all reactants (e.g. glucose
and oxygen) fortunately will not take place to any significant degree due to kinetic
barrieres (i.e. high activation energies, and no enzymes to give help) and so it can be bypassed; this allows the reaction to proceed via a series of intermediates, each with only
moderate –∆G´s, which can be better controlled and thus appropriately coupled with
formation of “energy rich” cofactors. b) “fuels” and intermediates generally are stable,
but cofactors might lose their enrgy by hydrolysis (ATP) or direct oxidation (NADH/
O2). At neutral pH hydrolysis of ATP is slow enough, but NADH-oxidation is a more
serious problem, that´s why the membrane electron transport system(s) have to be so
elaborate.