Abstract
Ferredoxin (Fd) in higher plants is encoded by a nuclear gene, synthesized in the cytoplasm as a larger precursor, and imported into the chloroplast, where it is proteolytically processed, and assembled with the [2Fe-2S] cluster. The final step in the biosynthetic pathway of Fd can be analyzed by a reconstitution system composed of isolated chloroplasts and [(35)S]cysteine, in which [(35)S]sulfide and iron are incorporated into Fd to build up the (35)S-labeled Fe-S cluster. Although a lysed chloroplast system shows obligate requirements for ATP and NADPH, in vitro chemical reconstitution of the Fe-S cluster is generally thought to be energy-independent. The present study investigated whether ATP and NADPH in the chloroplast system of spinach (Spinacia oleracea) are involved in the supply of [(35)S]sulfide or iron, or in Fe-S cluster formation itself. [(35)S]Sulfide was liberated from [(35)S] cysteine in an NADPH-dependent manner, whereas ATP was not necessary for this process. This desulfhydration of [(35)S]cysteine occurred before the formation of the (35)S-labeled Fe-S cluster, and the amount of radioactivity in [(35)S]sulfide was greater than that in (35)S-labeled holo-Fd by a factor of more than 20. Addition of nonradioactive sulfide (Na(2)S) inhibited competitively formation of the (35)S-labeled Fe-S cluster along with the addition of nonradioactive cysteine, indicating that some of the inorganic sulfide released from cysteine is incorporated into the Fe-S cluster of Fd. ATP hydrolysis was not involved in the production of inorganic sulfide or in the supply of iron for assembly into the Fe-S cluster. However, ATP-dependent Fe-S cluster formation was observed even in the presence of sufficient amounts of [(35)S]sulfide and iron. These results suggest a novel type of ATP-dependent in vivo Fe-S cluster formation that is distinct from in vitro chemical reconstitution. The implications of these results for the possible mechanisms of ATP-dependent Fe-S cluster formation are discussed.
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Claims 10
Claim · model-written summary
NADPH supported liberation of sulfide from cysteine.
- Taxon
- Spinach
Spinacia oleraceat0 - Treatment
- NADPHe0
- Outcome
- [(35)S]Sulfidee0
- Direction
- NADPH-dependente0
Not reportedComparator
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
t0“Spinach”
e0“[(35)S]Sulfide was liberated from [(35)S] cysteine in an NADPH-dependent manner, whereas ATP was not necessary for this process.”
Claim · model-written summary
ATP was unnecessary for sulfide liberation from cysteine.
- Taxon
- SpinachSpinacia oleraceat0
- Treatment
- ATPe0
- Outcome
- [(35)S]Sulfidee0
- Direction
- not necessarye0
Not reportedComparator
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
t0“Spinach”
e0“[(35)S]Sulfide was liberated from [(35)S] cysteine in an NADPH-dependent manner, whereas ATP was not necessary for this process.”
Claim · model-written summary
Cysteine desulfhydration preceded labeled Fe-S cluster formation.
- Taxon
- SpinachSpinacia oleraceat0
- Outcome
- desulfhydration of [(35)S]cysteinee0
- Qualifier
- before the formation of the (35)S-labeled Fe-S clustere0
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
t0“Spinach”
e0“This desulfhydration of [(35)S]cysteine occurred before the formation of the (35)S-labeled Fe-S cluster”
Claim · model-written summary
Radioactivity in sulfide exceeded that in holo-Fd by more than a factor of 20.
- Taxon
- SpinachSpinacia oleraceat0
- Outcome
- amount of radioactivity in [(35)S]sulfidee0
- Direction
- greater thane0
- Value
- more than 20e0
- Comparator
- (35)S-labeled holo-Fde0
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
t0“Spinach”
e0“the amount of radioactivity in [(35)S]sulfide was greater than that in (35)S-labeled holo-Fd by a factor of more than 20”
Claim · model-written summary
Nonradioactive sulfide competitively inhibited labeled Fe-S cluster formation.
- Taxon
- SpinachSpinacia oleraceat0
- Treatment
- nonradioactive sulfide (Na(2)S)e0
- Outcome
- formation of the (35)S-labeled Fe-S clustere0
- Direction
- inhibitede0
Not reportedComparator
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
t0“Spinach”
e0“Addition of nonradioactive sulfide (Na(2)S) inhibited competitively formation of the (35)S-labeled Fe-S cluster”
Claim · model-written summary
Nonradioactive cysteine competitively inhibited labeled Fe-S cluster formation.
- Taxon
- SpinachSpinacia oleraceat0
- Treatment
- nonradioactive cysteinee0
- Outcome
- formation of the (35)S-labeled Fe-S clustere0
- Direction
- inhibitede0
Not reportedComparator
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
t0“Spinach”
e0“Addition of nonradioactive sulfide (Na(2)S) inhibited competitively formation of the (35)S-labeled Fe-S cluster along with the addition of nonradioactive cysteine”
Claim · model-written summary
ATP hydrolysis was not involved in inorganic sulfide production.
- Taxon
- SpinachSpinacia oleraceat0
- Outcome
- production of inorganic sulfidee0
- Direction
- not involvede0
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
t0“Spinach”
e0“ATP hydrolysis was not involved in the production of inorganic sulfide or in the supply of iron for assembly into the Fe-S cluster”
Claim · model-written summary
ATP hydrolysis was not involved in iron supply for Fe-S cluster assembly.
- Taxon
- SpinachSpinacia oleraceat0
- Outcome
- supply of iron for assembly into the Fe-S clustere0
- Direction
- not involvede0
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
t0“Spinach”
e0“ATP hydrolysis was not involved in the production of inorganic sulfide or in the supply of iron for assembly into the Fe-S cluster”
Claim · model-written summary
Fe-S cluster formation remained ATP dependent despite sufficient sulfide and iron.
- Taxon
- SpinachSpinacia oleraceat0
- Outcome
- ATP-dependent Fe-S cluster formatione0
- Qualifier
- sufficient amounts of [(35)S]sulfide and irone0
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
t0“Spinach”
e0“ATP-dependent Fe-S cluster formation was observed even in the presence of sufficient amounts of [(35)S]sulfide and iron”
Claim · model-written summary
Some inorganic sulfide released from cysteine is incorporated into ferredoxin Fe-S clusters.
- Taxon
- SpinachSpinacia oleraceat0
- Compound
- inorganic sulfidee0
- Outcome
- Fe-S cluster of Fde0
Organisms pictured · Representative pictures of the matched species, not figures from the paper.
Verbatim
t0“Spinach”
e0“some of the inorganic sulfide released from cysteine is incorporated into the Fe-S cluster of Fd”
The same questions in other papers
ATP · molecular: metabolites · Spinacia oleracea (spinach)Positive and negative results
1 paper in the same direction as this paper · 2 in the opposite direction
- positive
- positive
- no effect
sulfide · molecular: metabolites · Spinacia oleracea (spinach)Results agree
1 paper in the same direction as this paper
- negative
NADPH · molecular: metabolites · Spinacia oleracea (spinach)Results agree
1 paper in the same direction as this paper
- positive
Claims are grouped by organism, treatment and what was measured or targeted. A paper counts as positive when any of its claims in the group is positive.