From the The PSI-C subunit of photosystem I (PS I) shows
similarity to soluble 2[4Fe-4S] ferredoxins. PSI-C contains an eight
residue internal loop and a 15 residue C-terminal extension which are absent in the ferredoxins. The eight-residue loop has been shown to
interact with PSI-A/PSI-B (Naver, H., Scott, M. P., Golbeck, J. H., Møller, B. L., and Scheller, H. V. (1996)
J. Biol. Chem. 271, 8996-9001). Four mutant proteins
were constructed. Two were modified barley PSI-C proteins, one lacking
the loop and the C terminus (PSI-Ccore) and one where the
loop replace the C-terminal extension
(PSI-CcoreLc-term). Two were modified
Clostridium pasteurianum ferredoxins, one with the loop of
barley PSI-C and one with both the loop and the C terminus of PSI-C.
Wild-type proteins and the mutants were used to reconstitute barley
P700-FX cores lacking PSI-C, -D, and-E. Western blotting
showed that PSI-CcoreLc-term binds to PS I,
whereas PSI-Ccore does not. Without PSI-D the
PSI-CcoreLc-term mutant accepts electrons from
FX in contrast to PSI-C mutants without the loop. Flash
photolysis of P700-FX cores reconstituted with C. pasteurianum ferredoxin showed that only the ferredoxin mutants
with the loop accepted electrons from FX. From this, it is
concluded that the loop of PSI-C is necessary and sufficient for the
association between PS I and PSI-C, and that the loop is functional as
an interaction domain even when positioned at the C terminus of PSI-C
or on a low molecular mass, soluble ferredoxin.
The photosystem I (PS
I)1 reaction center complex
mediates electron transport from plastocyanin to ferredoxin in oxygenic
photosynthesis. PS I contains the primary electron donor P700 (a Chl
a dimer) and the electron acceptors A0 (Chl
a), A1 (phylloquinone), and three
[4Fe-4S] centers FX, FA, and
FB (1-3). The terminal electron acceptors FA
and FB are bound to the PSI-C subunit (4, 5) while the
remaining electron acceptors are bound to the PSI-A/PSI-B heterodimer
(6, 7). Little is known about the nature of the interaction between
PSI-C and PSI-A/B. PSI-C interacts with two other extrinsic proteins
located at the stromal side of PS I, PSI-D, and PSI-E. In the absence
of PSI-D, PSI-C can still interact with PS I and accept electrons, but
stable binding of PSI-C is possible only when PSI-D is present (8, 9).
Furthermore, PSI-D is required for the docking of ferredoxin (10). The
presence or absence of PSI-E does not influence binding of PSI-C or
electron transport to FA and FB. PSI-E is
required for optimal rates of ferredoxin reduction but is not essential
(8, 11, 12).
The amino acid sequence of PSI-C is highly conserved among species and
contains two CXXCXXCXXXCP motifs which
are characteristic of 2[4Fe-4S] ferredoxins. The eight conserved
cysteine residues are ligands to the two [4Fe-4S] clusters in PSI-C
and ferredoxin and the conservation of the cysteine motifs shows that
the folding of PSI-C is similar to that of low molecular mass, soluble
iron-sulfur proteins whose three-dimensional structures have been
solved (e.g. Peptostreptococcus asaccharolyticus
ferredoxin) (13, 14). Apart from the cysteine motifs, PSI-C and the
ferredoxins show only little sequence similarity. Nevertheless,
alignment analysis clearly reveals two domains present only in PSI-C,
an internal segment of eight residues and a C-terminal addition of 15 residues. The published models of PSI-C show the eight additional
residues forming a loop extending from the known structure of P. asaccharolyticus ferredoxin (5, 15-17). In a previous study, we
reported that the eight-residue loop and the C-terminal addition play a
role in the interaction of PSI-C with PSI-A/B and PSI-D, respectively (18). These data have been supported by data of Rodday et
al. (19), who identified the two negatively charged residues
Glu-26 and Asp-31 of the loop as important for the PSI-C-PSI-A/B
interaction. In the present study, we show that the eight-residue loop
is necessary and sufficient for the functional association of
Clostridium pasteurianum ferredoxin with P700-FX
cores. The C-terminal domain mediates interaction with PSI-D but is not
sufficient for stable binding to the cores.
Using a modification of the method of Parrett et al. (20),
we showed that urea treatment of barley PS I complexes followed by a
combined detergent, salt, and urea wash leads to specific dissociation
of the PSI-C, -D, and -E subunits without affecting the remaining
polypeptides (8). The resulting P700-FX core can be used
for reconstitution by addition of Escherichia coli-expressed PSI-C and -D in the presence of the reagents Na2S,
FeCl3, and 2-mercaptoethanol, which serve to rebuild the
FeS clusters of PSI-C (8, 9, 21). In this work, we report the
specificity of the loop for interaction of PSI-C with the PSI-A/B
heterodimer by producing four modified proteins. Two were derived from
PSI-C, one lacking both the loop and the C-terminal extension, and one in which the loop replaced the residues in the C-terminal extension. Two were derived from soluble C. pasteurianum ferredoxin,
one in which the eight-residue loop from barley PSI-C was inserted between the two CXXCXXCXXXCP motifs
and one which contained both the loop and the C-terminal extension at
positions analogous to their locations in PSI-C. Using the in
vitro reconstitution system, we established that the eight-residue
internal loop of PSI-C is sufficient for interaction with the PS I
complex.
Plasmid Construction and Protein Isolation--
The production
of the PSI-C expression clone from barley, PSI-C Plant Biochemistry Laboratory,
Department of Biochemistry and Molecular
Biology, The Pennsylvania State University, University Park,
Pennsylvania 16802, and § Department of Biochemistry,
University of Nebraska-Lincoln, Lincoln, Nebraska 68588
ABSTRACT
Top
Abstract
Introduction
Procedures
Results
Discussion
References
INTRODUCTION
Top
Abstract
Introduction
Procedures
Results
Discussion
References
EXPERIMENTAL PROCEDURES
Top
Abstract
Introduction
Procedures
Results
Discussion
References
L and PSI-C
C has
been described previously (8, 18). PSI-C
C was derived from
Synechococcus sp. PCC 6301 psaC (22). However,
previous studies have shown that barley PSI-C and
Synechococcus sp. PCC 6301 PSI-C behave identically in
reconstitution experiments when using a barley P700-FX core
(18). The expression clone for C. pasteurianum ferredoxin
(23) was a gift from Dr. A. Wedd, University of Melbourne, Australia.
The amino acid sequences of the PSI-C mutants are shown in Fig.
1 where the amino acid sequence of the
ferredoxin from C. pasteurianum is compared with that of PSI-C from barley. To produce barley PSI-C lacking both the loop and
the C-terminal domain, appropriate oligonucleotides were used as
primers in a polymerase chain reaction with psaC
L as
template to yield one fragment. The psaCcore
gene was cloned into an NcoI and BamHI digested
Pet3d expression vector (Novagen, Stockholm, Sweden). PSI-C with the
loop at the C terminus was made by a polymerase chain reaction where
oligonucleotides were used as primers with psaC
L as
template and the resulting
psaCcoreLc-term gene was inserted
into Pet3d. To produce the C. pasteurianum ferredoxin (Cl.Fd) with the loop inserted, four oligonucleotides containing the
loop from PSI-C, were used with the Cl.Fd gene in two
polymerase chain reactions to yield the two products: the 5' end of the
gene with the whole loop and the 3' end with the whole loop. The two products which overlap in the loop region were used together in an
overlap extension polymerase chain reaction yielding one product, the
Cl.FdL gene. This being flanked by BamHI and
SalI sites was cloned into the BamHI and
SalI sites of the pGex-5x-3 vector (Pharmacia, Sollentuna,
Sweden). A polymerase chain reaction in which the 3' oligo encoded the
10 C-terminal residues of PSI-C in addition to the 3' end of
Cl.Fd was used to produce the Cl.FdLC gene, which was inserted into the BamHI and SalI sites of the
pGex-5x-3 vector. The phrase "the PSI-C polypeptides" is used for
the total of PSI-C, PSI-C
L,
PSI-CcoreLc-term, PSI-Ccore, and
PSI-C
C. The phrase "the clostridial ferredoxin polypeptides" is
used for the total of Cl.Fd, Cl.FdL, and Cl.FdLC. The PSI-D expression
clone from barley was made as described by Scott et al.
(25).
View larger version (43K):
[in a new window]
Fig. 1.
Amino acid sequences of mutant and wild-type
PSI-C (16) and C. pasteurianum ferredoxins (24). The
inserted loop sequence is shown and the applied C terminus is
underlined. The conserved cysteines in the
CXXCXXCXXXCP motifs are shown enlarged
in italics. Authentic barley PSI-C has serine as the
N-terminal residue rather than the methionine present in the
recombinant protein.
Mass Spectroscopy-- Matrix-assisted laser desorption ionization time-of-flight mass spectra of Cl.FdL and Cl.FdLC were recorded on a TofSpec E spectrometer (Micromass, Manchester, UK). Targets were prepared by loading 1 µl of a mixture of equal volumes sample and matrix solution (saturated sinapinic acid in 0.1% trifluoroacetic acid/MeCN, 6:4). In order to remove phosphate impurities, the air-dried target was washed with 1.5 µl of water and again allowed to dry. Bovine insulin and cytochrome c were added as internal standards.
Preparation of the P700-FX Core-- The PS I complex lacking PSI-C, PSI-D, and PSI-E from barley (Hordeum vulgare cv. Svalöfs Bonus) was isolated by a modification of the procedure of Andersen et al. (32) as described previously (18).
Rebinding of PSI-C, Cl.Fd, and PSI-D to the P700-FX
Core--
Reconstitutions were performed in 2-ml reaction volumes
containing 10 µg Chl/ml with a molar ratio of 20 PSI-C proteins or clostridial ferredoxin proteins per P700 according to Parrett et
al. (33) and Naver et al. (8, 18). When present, PSI-D was also used in a molar ratio of 20 per P700. The molar ratios of the
PS I subunits to P700 were determined by comparing the intensity of
Coomassie Brilliant Blue staining of SDS gels with a dilution series of
isolated subunits and known amounts of PS I. Molar ratios of the
clostridial ferredoxin proteins were determined spectroscopically after
rebuilding the iron-sulfur clusters using the extinction coefficient,
400 nm = 31.6 mM
1
cm
1 determined for Cl.Fd. After incubation for 16 h
at 4 °C the reconstituted complexes were used for the first set of
analyses. Subsequently, the samples were washed by ultrafiltration five
times over a YM 100 membrane (Amicon) using 50 mM Tris (pH
8.3), 0.1% Triton X-100 and used for EPR, NADP+
photoreduction or flash photolysis analysis.
EPR Spectroscopy-- EPR spectra were acquired as described previously (18). Illumination in the EPR cavity was performed by directing the beam of a 150-watt xenon arc lamp to the frozen sample through the slotted opening in the resonator. Photoaccumulation of electrons in the terminal acceptors was accomplished by passing the thawed sample through the focused beam of a 300-watt tungsten lamp into a container with liquid nitrogen over the course of 1 min.
Flash-induced Absorption Change--
Flash-induced absorption
transients at 834 nm were measured in a reaction mixture of 200-400
µl containing 50 mM Tris (pH 8.3), 2 mM
sodium ascorbate, 60 µM 2,6-dichlorophenolindophenol, and 10-20 µg
of Chl/ml using the setup described previously (18, 34). Reconstitution
is expressed as the percentage of the total absorbance change at 834 nm
that decayed with the long time constant (>30 ms) characteristic of
the P700+ [FA/FB]
back reaction.
NADP+ Photoreduction Measurements-- NADP+ photoreduction activity was determined from the absorbance change at 340 nm as described previously (18). NADP+ photoreduction measurements of the samples containing the clostridial ferredoxin proteins were carried out under strictly anaerobic conditions maintained using the glucose oxidase/catalase system as described by Kjaer and Scheller (35).
Additional Analytical Procedures-- Chlorophyll was determined according to Arnon (36). SDS-polyacrylamide gel electrophoresis was carried out in 8 to 25% gradient gels according to Fling and Gregerson (37). Prior to electrophoresis, the reconstituted PS I complexes were pelleted by ultracentrifugation. Western blot analysis was carried out by transferring electrophoresed proteins to nitrocellulose membranes followed by incubation with monospecific rabbit antibodies (32) and visualized using a secondary antibody conjugated with alkaline phosphatase (DAKO, Copenhagen, Denmark).
![]() |
RESULTS |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
Characterization of the Rebinding of PSI-C to the
P700-FX Core by Western Blot Analysis--
Treatment with
urea leads to dissociation of the PSI-C subunit from barley PS I. Reconstitution using the PSI-C polypeptides and PSI-D resulted in
rebinding of each of PSI-C, PSI-CL, PSI-C
C, and
PSI-CcoreLc-term as shown by Western blot
analysis of the washed complexes (Fig.
2). The PSI-Ccore did not
bind to the P700-FX core. As expected the PSI-C
C and the
PSI-Ccore migrate faster in SDS-polyacrylamide gel
electrophoresis than PSI-C. The PSI-C
L and
PSI-CcoreLc-term migrate the same as the
full-length PSI-C; however, the amino acid sequence of PSI-C
L has
been confirmed by protein sequencing (18). The band around 18 kDa seen
in some lanes probably represents a small amount of PSI-C dimers
present due to insufficient solubilization.
|
Flash-induced Absorbance Changes of P700-FX Cores
Reconstituted with the PSI-C Polypeptides--
The back reactions from
the PS I electron acceptors were measured by flash induced absorbance
changes at 834 nm after addition of each of the PSI-C polypeptides to
P700-FX cores in the presence and absence of PSI-D. In a
P700-FX core where FA and FB are
absent, the charge recombination between FX
and P700+ will proceed with a time constant of
approximately 1 ms (38). In contrast, when iron-sulfur clusters
FA and FB are functional, the forward electron
transfer from FX
to
[FA/FB] is so efficient that no back reaction
from FX
can take place and only the much
slower (>30 ms) back reaction from
[FA/FB]
is observed. Thus the
degree of functional reconstitution can be determined by observing the
replacement of the
1-ms back reaction of P700+
FX
with the slower back reaction
P700+[FA/FB]
Addition of PSI-C
L and PSI-CcoreLc-term to
P700-FX cores in the absence of PSI-D resulted in 12 and
60% recovery of the slow phase, respectively (Fig.
3, Table
I), indicating the importance of the loop
sequence for electron transport. In contrast, when PSI-D was present
reconstitution with PSI-C
L resulted in a substantial recovery of the
slow phase (52%) while reconstitution with
PSI-CcoreLc-term resulted in an improved
recovery of the slow phase (73%). The PSI-Ccore did not
show any restoration with the P700-FX core in the presence
or absence of PSI-D. To assay the stability of the associations, the
reaction mixtures were washed with 0.1% Triton X-100 over an
ultrafiltration membrane. As expected from earlier studies with
wild-type PSI-C (39), none of the associations was stable in the
absence of PSI-D (Table I). In the presence of PSI-D the associations
were stable, although a small decrease in the slow phase was observed
with all the PSI-C proteins except the wild-type protein.
|
|
EPR Analysis of the PS I Complexes Reconstituted with PSI-D and PSI-CcoreLc-term-- Iron-sulfur clusters can be reconstituted in all of the PSI-C polypeptides as shown by their absorbance spectra (26) (data not shown). Having found that PSI-CcoreLc-term binds to PS I and accepts electrons from FX, the next task was to determine if the EPR properties of FA and FB were different from those of wild-type PSI-C. The reconstituted PS I complex was illuminated at 15 K in the EPR cavity to promote one electron to the acceptor system. The resulting spectrum showed that the field positions, line widths and ratios of FB (g = 2.065 and 1.931) and FA (g = 2.048, 1.947, and 1.869) were identical to that of a P700-FX core reconstituted with PSI-C and PSI-D (Fig. 4A). The reconstituted PS I complex was next frozen during illumination, a protocol which allows two or more electrons to accumulate in the electron acceptor system. The resulting interaction spectrum was similar to that of a wild-type PSI-C reconstitution, containing the features characteristic of magnetic coupling between FA and FB (Fig. 4B).
|
Reconstitution of the P700-FX Core Using the
Clostridial Ferredoxin Proteins--
The pure clostridial ferredoxin
proteins were obtained after cleavage of the fusion proteins by factor
Xa. The amount of holo protein obtained after rebuilding the
iron-sulfur clusters was determined by UV-visible spectroscopy using
400 nm = 31.6 mM
1
cm
1. The UV-visible difference spectra of mutant proteins
Cl.FdL and Cl.FdLC resembles that of PSI-C (Fig.
5). The molecular masses of Cl.FdL and
Cl.FdLC were determined by matrix-assisted laser desorption ionization
time-of-flight spectrometry to 6808.8 and 7947.4 Da, respectively.
These values are within 0.3-0.6 mass units of the calculated molecular
masses of 6806.4 and 7946.7 Da. It is therefore not likely that any
secondary modification has taken place during the expression of the
proteins.
|
|
|
![]() |
DISCUSSION |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|
The removal of both the loop and the C-terminal extension from
PSI-C led to a protein (PSI-Ccore) unable to bind to PS I
or accept electrons from FX. Thus, to obtain a functional
PSI-C it is essential that either the loop domain and/or the C-terminal domain is present. PSI-CL is unable to accept electrons from FX when PSI-D is not present. However, when the C-terminal
extension of this mutant was replaced with the loop sequence, the
resulting protein (PSI-CcoreLc-term) is able to
accept electrons from FX, even when PSI-D is not present.
Therefore the interaction between PSI-C and PSI-A/B must depend only on
the presence of the loop sequence, independent of its position in the
protein. Since the orientation of the functional PSI-C protein has been
limited to two possibilities (14), and since the loop and C terminus of PSI-C have been predicted to extend from opposite sides of the molecule
(5, 15-17) it is possible that the PS
I-PSI-CcoreLc-term interaction involves an
inverted orientation of the PSI-CcoreLc-term compared with the wild type PSI-C. The EPR measurements would seem to
contradict this notion since the spectra of the P700-FX core reconstituted with wild type PSI-C and with
PSI-CcoreLc-term are almost identical. We can
envision two mechanisms to explain these results. Illumination of PS I
at cryogenic temperatures result in reduction of FA in 76%
and FB in 24% of the complexes (40). This distribution is
similar to that calculated assuming an equilibrium between
FB and FA with redox midpoint potentials at
580 mV and
520 mV, respectively (41). If the reduction potentials
of FA and FB are not altered by misbinding to
PS I (40), and if the electron equilibrates between the two acceptors at low temperature, then the same charge distribution should be expected even if the mutated PSI-C protein were oriented differently than the wild-type. Alternatively, the resonances of the FA
and FB clusters in the mutant protein may be transposed.
Studies of unbound mutant PSI-C proteins show that the EPR resonances
of FA and FB are virtually indistinguishable in
terms of g values and line widths (36). When the mutant
PSI-C proteins are bound to PS I, the FA and FB
resonances narrow and become differentiated with respect to their
principal g values (42, 43). Thus, the final determinants of
the g tensors of FA and FB may be
ultimately related to protein:protein interactions with PSI-A/B and
PSI-D (41). It is important to note that the PS I complex reconstituted with the PSI-CcoreLc-term does not support
NADP+ reduction. Apparently, those determinants which
confer binding and/or electron transfer properties from
FA/FB to ferredoxin are more stringent than
those which confer forward electron transfer from FX to
FA/FB. EPR analysis after chemical reduction in
the dark showed that the reduction potential of
PSI-CcoreLc-term is not significantly higher
than that of the wild-type PSI-C (data not shown). Thus, the inability
to reduce ferredoxin efficiently is a consequence of improper docking
of ferredoxin rather than of a change in the equilibrium of the redox
reaction. The analysis of the two mutants PSI-Ccore and
PSI-CcoreLc-term therefore lead to the
following conclusions: 1) the loop residues of PSI-C interact specifically with PSI-A/B even when it is moved to the C terminus of
the PSI-C and 2) The binding of PSI-C to PS I is carried out by the two
domains, the loop and the C terminus since deletion of both results in
no interaction between the mutant and PS I.
The Cl.Fd protein is unable to bind to PS I or accept electrons from
FX, however when the loop residues of PSI-C were added, the
resulting protein Cl.FdL was able to accept electrons from FX. This result shows that the Cl.FdL and Cl.FdLC are in
fact associated with PS I although weakly. If the proteins merely
function as soluble electron acceptors and the electron transfer is a
diffusion limited process, it is unlikely that the second-order rate
constant would be high enough with submicromolar concentrations of
these proteins to out-compete the back reaction from FX.
The PS I complexes reconstituted with Cl.FdL or Cl.FdLC were not able
to support NADP+ reduction. Most likely the Cl.Fd proteins
do not interact properly with barley ferredoxin. One residue of PSI-C,
namely lysine in position 34, has been identified as essential for the
PSI-C-ferredoxin interaction (44). Cl.Fd does not have a lysine or any
equivalent around position 34. Another factor may be that Cl.Fd does
not have sufficiently low midpoint potential to reduce ferredoxin with
sufficient efficiency to out-compete the back reaction from FX. Cl.Fd has a redox potential of 420 mV (45) which is
similar to that of plant [2Fe-2S] ferredoxin (
401 mV) (46), while
FA has a redox potential of
520 mV (41).
Cl.Fd is a far more acidic protein than PSI-C, but the addition of the
eight residues of the loop in PSI-C to Cl.Fd is apparently sufficient
to allow the protein (Cl.FdL) to interact with PS I. The addition of
the C terminus of PSI-C to Cl.FdL produced a protein Cl.FdLC which
gives higher recovery of the slow rereduction in reconstitution
experiments than Cl.FdL, but only when PSI-D was present. Cl.FdL
appears to interact with the P700-FX core to the same
extent in the presence and absence of PSI-D. A similar result was
obtained previously for the PSI-CC mutant (18) (Table I), and these
results support the earlier conclusion that the C-terminal extension of
PSI-C provides an interaction site for PSI-D. The Cl.FdLC protein
accepted electrons from FX significantly more efficiently
than the Cl.FdL protein electrons from FX and, by inference, interact with PSI-D. However, the association of Cl.FdLC was
not sufficiently stable to survive a washing step. Reconstitution with
PSI-CcoreLc-term and PSI-C
C both show a
higher degree of stability in the presence of PSI-D despite the lack of
the C terminus. These results show that PSI-C contains residues apart
from the C terminus which are also important for the interaction with
PSI-D. Further site-specific mutagenesis will be required to identify these sites of interaction with PSI-D. Contrary to this result, the
loop affects binding of PSI-C and Cl.Fd in a very similar manner. Thus,
residues of PSI-C other than the eight residue internal loop appear to
be of little consequence for the binding to the P700-FX
core itself. Recent data of Rodday et al. (19) reveal that
not only the two acidic residues of the loop (residues 26 and 31) but
also the aspartic acid residue 8 are important for the interaction.
However, this aspartic acid is conserved among soluble 2[4Fe-4S]
ferredoxins and thus also present in the Clostridium ferredoxin (Fig. 1).
The interaction between the loop and the PSI-A/B heterodimer is most simply understood if PSI-C is placed on PSI-A/B with the iron-sulfur cluster FB proximal to FX as previously proposed, since in this orientation the loop would face the membrane (1, 14, 47, 48). In contrast, the inability for electron transfer to occur to ferredoxin in FB-deficient PS I complexes and the resumption of electron transfer to ferredoxin in FB-reconstituted PS I complexes favor the opposite orientation of PSI-C with FA proximal to FX (49-51) (see also discussion in Brettel (48)). In this orientation the loop would seem to extend from a position on PSI-C facing the stroma. However, we note that helices j/j' and k/k' in the FX-binding region of PSI-A and PSI-B are connected with loops of about 40 amino acid residues (52). These loops are poorly resolved in the electron density map and could extend a considerable distance away from the membrane and contact the PSI-C loop in both suggested orientations of PSI-C. The resolution of the issue of PSI-C orientation ultimately awaits an x-ray crystal structure at a higher level of resolution.
In conclusion, the eight-residue loop of PSI-C is essential for the interaction of PSI-C and PS I. The domain is functional in mediating protein-protein association both when placed as the C terminus of the PSI-C or when inserted into Clostridium ferredoxin between the two CXXCXXCXXXCP motifs.
![]() |
ACKNOWLEDGEMENTS |
---|
We thank Hanne Linde Nielsen and Inga Olsen for technical assistance and Dr. A. Wedd for the C. pasteurianum ferredoxin expression clone. Dr. Birger Lindberg Møller and Dr. Norbert Krauss are thanked for valuable discussions.
![]() |
FOOTNOTES |
---|
* This work was supported in part by grants from the Danish Center for Plant Biotechnology, The Danish National Science Research Council, and the National Science Foundation (MCB-9205756).The costs of publication of this article were defrayed in part by the payment of page charges. The article must therefore be hereby marked "advertisement" in accordance with 18 U.S.C. Section 1734 solely to indicate this fact.
¶ Present address: USDA-ARS, Dept. of Agronomy, Iowa State University, Ames, IA 50011.
To whom correspondence should be addressed. Tel.: 45 35283354, Fax: 45 35283333, E-mail: scheller{at}biobase.dk.
1
The abbreviations used are: PS I, photosystem I;
Chl, chlorophyll; Cl.Fd, C. pasteurianum ferredoxin; Cl.FdL,
C. pasteurianum ferredoxin with residue 26-33 of PSI-C from
barley inserted at residue 23; Cl.FdLC, C. pasteurianum
ferredoxin with residue 26-33 of PSI-C from barley inserted at residue
23 and with the 10 C-terminal residues of PSI-C applied as a C-terminal
extension; FNR, ferredoxin:NADP+ oxidoreductase, PSI-CC,
PSI-C from Synechococcus sp. 6301 lacking the 10 C-terminal
residues; PSI-C, PSI-C from barley; PSI-C
L, PSI-C from barley
lacking residues 26-33; PSI-Ccore, PSI-C from barley
lacking the 10 C-terminal residues and residue 26-33;
PSI-CcoreLc-term, PSI-C from barley where
residue 26-33 has been moved to the C terminus so that they replace
the original 10 C-terminal residues.
![]() |
REFERENCES |
---|
![]() ![]() ![]() ![]() ![]() ![]() ![]() |
---|