Fluorescence quenching of IsiA in early stage of iron deficiency and at cryogenic temperatures

Cyanobacteria respond to iron deficiency during growth by expressing the isiA gene, which produces a chlorophyll-carotenoid protein complex known as IsiA or CP43′. Long-term iron deficiency results in the formation of large IsiA aggregates, some of which associate with photosystem I (PSI) while others are not connected to a photosystem. The fluorescence at room temperature of these unconnected aggregates is strongly quenched, which points to a photoprotective function. In this study, we report time-resolved fluorescence measurements of IsiA aggregates at low temperatures. The average fluorescence lifetimes are estimated to be about 600 ps at 5 K and 150 ps at 80 K. Both lifetimes are much shorter than that of the monomeric complex CP47 at 77 K. We conclude that IsiA aggregates quench fluorescence to a significant extent at cryogenic temperatures. We show by low-temperature fluorescence spectroscopy that unconnected IsiA is present already after two days of growth in an iron-deficient medium, when PSI and PSII are still present in significant amounts and that under these conditions the fluorescence quenching is similar to that after 18 days, when PSI is almost completely absent. We conclude that unconnected IsiA provides photoprotection in all stages of iron deficiency.     

Aggregates of the chlorophyll-binding protein IsiA (CP43') dissipate energy in cyanobacteria

In many natural habitats, growth of cyanobacteria may be limited by a low concentration of iron. Cyanobacteria respond to this condition by expressing a number of iron-stress-inducible genes, of which the isiA gene encodes a chlorophyll-binding protein known as IsiA or CP43'. IsiA monomers assemble into ring-shaped polymers that encircle trimeric or monomeric photosystem I (PSI), or are present in supercomplexes without PSI, in particular upon prolonged iron starvation. In this report, we present steady-state and time-resolved fluorescence measurements of isolated IsiA aggregates that have been purified from an iron-starved psaFJ-minus mutant of Synechocystis PCC 6803. We show that these aggregates have a fluorescence quantum yield of approximately 2% compared to that of chlorophyll a in acetone, and that the dominating fluorescence lifetimes are 66 and 210 ps, more than 1 order of magnitude shorter than that of free chlorophyll a. Comparison of the temperature dependence of the fluorescence yields and spectra of the isolated aggregates and of the cells from which they were obtained suggests that these aggregates occur naturally in the iron-starved cells. We suggest that IsiA aggregates protect cyanobacterial cells against the deleterious effects of light.     

Spectroscopic properties of PSI-IsiA supercomplexes from the cyanobacterium Synechococcus PCC 7942

The cyanobacterium Synechococcus PCC 7942 grown under iron starvation assembles a supercomplex consisting of a trimeric Photosystem I (PSI) complex encircled by a ring of 18 CP43' or IsiA light-harvesting complexes [Nature 412 (2001) 745]. Here we present a spectroscopic characterization by temperature-dependent absorption and fluorescence spectroscopy, site-selective fluorescence spectroscopy at 5 K, and circular dichroism of isolated PSI-IsiA, PSI and IsiA complexes from this cyanobacterium grown under iron starvation. The results suggest that the IsiA ring increases the absorption cross-section of PSI by about 100%. Each IsiA subunit binds about 16-17 chlorophyll a (Chl a) molecules and serves as an efficient antenna for PSI. Each of the monomers of the trimeric PSI complex contains two red chlorophylls, which presumably give rise to one exciton-coupled dimer and at 5 K absorb and fluoresce at 703 and 713 nm, respectively. The spectral properties of these C-703 chlorophylls are not affected by the presence of the IsiA antenna ring. The spectroscopic properties of the purified IsiA complexes are similar to those of the related CP43 complex from plants, except that the characteristic narrow absorption band of CP43 at 682.5 nm is missing in IsiA.     

Supramolecular organization and dual function of the IsiA chlorophyll-binding protein in cyanobacteria

A significant part of global primary productivity is provided by cyanobacteria, which are abundant in most marine and freshwater habitats. In many oceanographic regions, however, the concentration of iron can be so low that it limits growth. Cyanobacteria respond to this condition by expressing a number of iron stress inducible genes, of which the isiA gene encodes a chlorophyll-binding protein known as IsiA or CP43'. It was recently shown that 18 IsiA proteins encircle trimeric photosystem I (PSI) under iron-deficient growth conditions. We report here that after prolonged growth of Synechocystis PCC 6803 in an iron-deficient medium, the number of bound IsiA proteins can be much higher than previously known. The largest complexes bind 12-14 units in an inner ring and 19-21 units in an outer ring around a PSI monomer. Fluorescence excitation spectra indicate an efficient light harvesting function for all PSI-bound chlorophylls. We also find that IsiA accumulates in cyanobacteria in excess of what is needed for functional light harvesting by PSI, and that a significant part of IsiA builds supercomplexes without PSI. Because the further decline of PSI makes photosystem II (PSII) increasingly vulnerable to photooxidation, we postulate that the surplus synthesis of IsiA shields PSII from excess light. We suggest that IsiA plays a surprisingly versatile role in cyanobacteria, by significantly enhancing the light harvesting ability of PSI and providing photoprotection for PSII.     

The identification of IsiA proteins binding chlorophyll <Emphasis Type="Italic">d</Emphasis> in the cyanobacterium <Emphasis Type="Italic">Acaryochloris marina</Emphasis>

The bioavailable iron in many aquatic ecosystems is extremely low, and limits the growth and photosynthetic activity of phytoplankton. In response to iron limitation, a group of chlorophyll-binding proteins known as iron stress-induced proteins are induced and serve as accessory light-harvesting components for photosystems under iron limitation. In the present study, we investigated physiological features of Acaryochloris marina in response to iron-deficient conditions. The growth doubling time under iron-deficient conditions was prolonged to ~3.4 days compared with 1.9 days under normal culture conditions, accompanied with dramatically decreased chlorophyll content. The isolation of chlorophyll-binding protein complexes using sucrose density gradient centrifugation shows six main green bands and three main fluorescence components of 712, 728, and 748 nm from the iron-deficient culture. The fluorescence components of 712 and 728 nm co-exist in the samples collected from iron-deficient and iron-replete cultures and are attributed to Chl d-binding accessory chlorophyll-binding antenna proteins and also from photosystem II. A new chlorophyll-binding protein complex with its main fluorescence peak at 748 nm was observed and enriched in the heaviest fraction from the samples collected from the iron-deficient culture only. Combining western blotting analysis using antibodies of CP47 (PSII), PsaC (PSI) and IsiA and proteomic analysis on an excised protein band at ~37 kDa, the heaviest fraction (?F6) isolated from iron-deficient culture contained Chl d-bound PSI–IsiA supercomplexes. The PSII-antenna supercomplexes isolated from iron-replete conditions showed two fluorescence peaks of 712 and 728 nm, which can be assigned as 6-transmembrane helix chlorophyll-binding antenna and photosystem II fluorescence, respectively, which is supported by protein analysis of the fractions (F5 and F6).     

Time-resolved absorption and emission show that the CP43' antenna ring of iron-stressed synechocystis sp. PCC6803 is efficiently coupled to the photosystem I reaction center core

Excitation energy transfer and trapping processes in an iron stress-induced supercomplex of photosystem I from the cyanobacterium Synechocystis sp. PCC6803 were studied by time-resolved absorption and fluorescence spectroscopy on femtosecond and picosecond time scales. The data provide evidence that the energy transfer dynamics of the CP43'-PSI supercomplex are consistent with energy transfer processes that occur in the Chl a network of the PSI trimer antenna. The most significant absorbance changes in the CP43'-PSI supercomplex are observed within the first several picoseconds after the excitation into the spectral region of CP43' absorption (665 nm). The difference time-resolved spectra (DeltaDeltaA) resulting from subtraction of the PSI trimer kinetic data from the CP43'-PSI supercomplex data indicate three energy transfer processes with time constants of 0.2, 1.7, and 10 ps. The 0.2 ps kinetic phase is tentatively interpreted as arising from energy transfer processes originating within or between the CP43' complexes. The 1.7 ps phase is interpreted as possibly arising from energy transfer from the CP43' ring to the PSI trimer via closely located clusters of Chl a in CP43' and the PSI core, while the slower 10 ps process might reflect the overall excitation transfer from the CP43' ring to the PSI trimer. These three fast kinetic phases are followed by a 40 ps overall excitation decay in the supercomplex, in contrast to a 25 ps overall decay observed in the trimer complex without CP43'. Excitation of Chl a in both the CP43'-PSI antenna supercomplex and the PSI trimer completely decays within 100 ps, resulting in the formation of P700(+). The data indicate that there is a rapid and efficient energy transfer between the outer antenna ring and the PSI reaction center complex.     

Light-induced energy dissipation in iron-starved cyanobacteria: roles of OCP and IsiA proteins

In response to iron deficiency, cyanobacteria synthesize the iron stress-induced chlorophyll binding protein IsiA. This protein protects cyanobacterial cells against iron stress. It has been proposed that the protective role of IsiA is related to a blue light-induced nonphotochemical fluorescence quenching (NPQ) mechanism. In iron-replete cyanobacterial cell cultures, strong blue light is known to induce a mechanism that dissipates excess absorbed energy in the phycobilisome, the extramembranal antenna of cyanobacteria. In this photoprotective mechanism, the soluble Orange Carotenoid Protein (OCP) plays an essential role. Here, we demonstrate that in iron-starved cells, blue light is unable to quench fluorescence in the absence of the phycobilisomes or the OCP. By contrast, the absence of IsiA does not affect the induction of fluorescence quenching or its recovery. We conclude that in cyanobacteria grown under iron starvation conditions, the blue light-induced nonphotochemical quenching involves the phycobilisome OCP-related energy dissipation mechanism and not IsiA. IsiA, however, does seem to protect the cells from the stress generated by iron starvation, initially by increasing the size of the photosystem I antenna. Subsequently, the IsiA converts the excess energy absorbed by the phycobilisomes into heat through a mechanism different from the dynamic and reversible light-induced NPQ processes.     

Excitation energy transfer in native and unstacked thylakoid membranes studied by low temperature and ultrafast fluorescence spectroscopy

In this work, the transfer of excitation energy was studied in native and cation-depletion induced, unstacked thylakoid membranes of spinach by steady-state and time-resolved fluorescence spectroscopy. Fluorescence emission spectra at 5 K show an increase in photosystem I (PSI) emission upon unstacking, which suggests an increase of its antenna size. Fluorescence excitation measurements at 77 K indicate that the increase of PSI emission upon unstacking is caused both by a direct spillover from the photosystem II (PSII) core antenna and by a functional association of light-harvesting complex II (LHCII) to PSI, which is most likely caused by the formation of LHCII-LHCI-PSI supercomplexes. Time-resolved fluorescence measurements, both at room temperature and at 77 K, reveal differences in the fluorescence decay kinetics of stacked and unstacked membranes. Energy transfer between LHCII and PSI is observed to take place within 25 ps at room temperature and within 38 ps at 77 K, consistent with the formation of LHCII-LHCI-PSI supercomplexes. At the 150–160 ps timescale, both energy transfer from LHCII to PSI as well as spillover from the core antenna of PSII to PSI is shown to occur at 77 K. At room temperature the spillover and energy transfer to PSI is less clear at the 150 ps timescale, because these processes compete with charge separation in the PSII reaction center, which also takes place at a timescale of about 150 ps.     

Iron deficiency in cyanobacteria causes monomerization of photosystem I trimers and reduces the capacity for state transitions and the effective absorption cross section of photosystem I in vivo

The induction of the isiA (CP43') protein in iron-stressed cyanobacteria is accompanied by the formation of a ring of 18 CP43' proteins around the photosystem I (PSI) trimer and is thought to increase the absorption cross section of PSI within the CP43'-PSI supercomplex. In contrast to these in vitro studies, our in vivo measurements failed to demonstrate any increase of the PSI absorption cross section in two strains (Synechococcus sp. PCC 7942 and Synechocystis sp. PCC 6803) of iron-stressed cells. We report that iron-stressed cells exhibited a reduced capacity for state transitions and limited dark reduction of the plastoquinone pool, which accounts for the increase in PSII-related 685 nm chlorophyll fluorescence under iron deficiency. This was accompanied by lower abundance of the NADP-dehydrogenase complex and the PSI-associated subunit PsaL, as well as a reduced amount of phosphatidylglycerol. Nondenaturating polyacrylamide gel electrophoresis separation of the chlorophyll-protein complexes indicated that the monomeric form of PSI is favored over the trimeric form of PSI under iron stress. Thus, we demonstrate that the induction of CP43' does not increase the PSI functional absorption cross section of whole cells in vivo, but rather, induces monomerization of PSI trimers and reduces the capacity for state transitions. We discuss the role of CP43' as an effective energy quencher to photoprotect PSII and PSI under unfavorable environmental conditions in cyanobacteria in vivo.     

The photochemical trapping rate from red spectral states in PSI-LHCI is determined by thermal activation of energy transfer to bulk chlorophylls

The average fluorescence decay lifetimes, due to reaction centre photochemical trapping, were calculated for wavelengths in the 690- to 770-nm interval from the published fluorescence decay-associated emission spectra for Photosystem I (PSI)-light-harvesting complex of Photosystem I (LHCI) [Biochemistry 39 (2000) 6341] at 280 and 170 K. For 280 K, the overall trapping time at 690 nm is 81 ps and increases with wavelength to reach 103 ps at 770 nm. For 170 K, the 690-nm value is 115 ps, increasing to 458 ps at 770 nm. This underlines the presence of kinetically limiting processes in the PSI antenna (diffusion limited). The explanation of these nonconstant values for the overall trapping time band is sought in terms of thermally activated transfer from the red absorbing states to the "bulk" acceptor chlorophyll (chl) states in the framework of the Arrhenius-Eyring theory. It is shown that the wavelength-dependent "activation energies" come out in the range between 1.35 and 2.7 kcal mol(-1), increasing with the emission wavelength within the interval 710-770 nm. These values are in good agreement with the Arrhenius activation energy determined for the steady-state fluorescence yield over the range 130-280 K for PSI-LHCI. We conclude that the variable trapping time in PSI-LHCI can be accounted for entirely by thermally activated transfer from the low-energy chl states to the bulk acceptor states and therefore that the position of the various red states in the PSI antenna seems not to be of significant importance. The analysis shows that the bulk antenna acceptor states are on the low-energy side of the bulk antenna absorption band.     

The photochemical trapping rate from red spectral states in PSI–LHCI is determined by thermal activation of energy transfer to bulk chlorophylls

The average fluorescence decay lifetimes, due to reaction centre photochemical trapping, were calculated for wavelengths in the 690- to 770-nm interval from the published fluorescence decay-associated emission spectra for Photosystem I (PSI)–light-harvesting complex of Photosystem I (LHCI) [Biochemistry 39 (2000) 6341] at 280 and 170 K. For 280 K, the overall trapping time at 690 nm is 81 ps and increases with wavelength to reach 103 ps at 770 nm. For 170 K, the 690-nm value is 115 ps, increasing to 458 ps at 770 nm. This underlines the presence of kinetically limiting processes in the PSI antenna (diffusion limited). The explanation of these nonconstant values for the overall trapping time band is sought in terms of thermally activated transfer from the red absorbing states to the “bulk” acceptor chlorophyll (chl) states in the framework of the Arrhenius–Eyring theory. It is shown that the wavelength-dependent “activation energies” come out in the range between 1.35 and 2.7 kcal mol⁻¹, increasing with the emission wavelength within the interval 710–770 nm. These values are in good agreement with the Arrhenius activation energy determined for the steady-state fluorescence yield over the range 130–280 K for PSI–LHCI. We conclude that the variable trapping time in PSI–LHCI can be accounted for entirely by thermally activated transfer from the low-energy chl states to the bulk acceptor states and therefore that the position of the various red states in the PSI antenna seems not to be of significant importance. The analysis shows that the bulk antenna acceptor states are on the low-energy side of the bulk antenna absorption band.     

Photosystem I trimers from Synechocystis PCC 6803 lacking the PsaF and PsaJ subunits bind an IsiA ring of 17 units

We report a structural characterization by electron microscopy and image analysis of a supramolecular complex consisting of Photosystem I (PSI) and the chlorophyll-binding protein IsiA from a mutant of the cyanobacterium Synechocystis PCC 6803 lacking the PsaF and PsaJ subunits. The circular complex consists of a central PSI trimer surrounded by a ring of 17 IsiA units, one less than in the wild-type supercomplex. We conclude that PsaF and PsaJ are not obligatory for the binding of the IsiA ring, and that the size of the PSI complex determines the number of IsiA units in the ring. The resulting number of 17 copies implies that each PSI monomer has a different association to the IsiA ring.     

Pathways for energy transfer in the core light-harvesting complexes CP43 and CP47 of photosystem II

The pigment-protein complexes CP43 and CP47 transfer excitation energy from the peripheral antenna of photosystem II toward the photochemical reaction center. We measured the excitation dynamics of the chlorophylls in isolated CP43 and CP47 complexes at 77 K by time-resolved absorbance-difference and fluorescence spectroscopy. The spectral relaxation appeared to occur with rates of 0.2-0.4 ps and 2-3 ps in both complexes, whereas an additional relaxation of 17 ps was observed only in CP47. Using the 3.8-A crystal structure of the photosystem II core complex from Synechococcus elongatus (A. Zouni, H.-T. Witt, J. Kern, P. Fromme, N. Krauss, W. Saenger, and P. Orth, 2001, Nature, 409:739-743), excitation energy transfer kinetics were calculated and a Monte Carlo simulation of the absorption spectra was performed. In both complexes, the rate of 0.2-0.4 ps can be ascribed to excitation energy transfer within a layer of chlorophylls near the stromal side of the membrane, and the slower 2-3-ps process to excitation energy transfer to the calculated lowest excitonic state. We conclude that excitation energy transfer within CP43 and CP47 is fast and does not contribute significantly to the well-known slow trapping of excitation energy in photosystem II.     

Photosystem I trimers from Synechocystis PCC 6803 lacking the PsaF and PsaJ subunits bind an IsiA ring of 17 units

We report a structural characterization by electron microscopy and image analysis of a supramolecular complex consisting of Photosystem I (PSI) and the chlorophyll-binding protein IsiA from a mutant of the cyanobacterium Synechocystis PCC 6803 lacking the PsaF and PsaJ subunits. The circular complex consists of a central PSI trimer surrounded by a ring of 17 IsiA units, one less than in the wild-type supercomplex. We conclude that PsaF and PsaJ are not obligatory for the binding of the IsiA ring, and that the size of the PSI complex determines the number of IsiA units in the ring. The resulting number of 17 copies implies that each PSI monomer has a different association to the IsiA ring.     

Mobility of the IsiA chlorophyll-binding protein in cyanobacterial thylakoid membranes

We are using fluorescence recovery after photobleaching (FRAP) to probe the dynamics of thylakoid membranes in vivo in cells of the cyanobacterium Synechococcus sp. PCC7942. We have shown previously that the light-harvesting phycobilisomes diffuse quite rapidly on the thylakoid membrane surface. However, the photosystem II core complexes appear completely immobile. This raises the possibility that all of the membrane integral protein complexes in the thylakoid membrane are locked into a rather rigid array. Alternatively, it is possible that photosystem II is specifically anchored in the membrane, with other membrane proteins able to diffuse around it. We have now resolved this question by studying the diffusion of a second integral membrane protein, the IsiA chlorophyll-binding protein. IsiA is induced under iron starvation and some other stress conditions. In iron-stressed cyanobacterial cells, a high proportion of chlorophyll fluorescence comes from IsiA. This makes it straightforward to examine the diffusion of IsiA by FRAP. We find that the complex is mobile with a mean diffusion coefficient of approximately 3 x 10(-11) cm(2) s(-1). Thus it is clear that some thylakoid membrane proteins are mobile and that there must be a specific anchor that prevents photosystem II diffusion. We discuss the implications for the structure and function of the cyanobacterial thylakoid membrane.     

Ferritin is required for rapid remodeling of the photosynthetic apparatus and minimizes photo-oxidative stress in response to iron availability in Chlamydomonas reinhardtii

Ferritin is a key player in the iron homeostasis due to its ability to store large quantities of iron. Chlamydomonas reinhardtii contains two nuclear genes for ferritin ( ferr1 and ferr2 ) that are induced when Chlamydomonas cells are shifted to iron-deficient conditions. In response to the reduced iron availability, degradation of photosystem I (PSI) and remodeling of its light-harvesting complex occur. This active PSI degradation slows down under photo-autotrophic conditions where photosynthesis is indispensable. We observed a strong induction of ferritin correlated with the degree of PSI degradation during iron deficiency. The PSI level can be restored to normal within 24 h after iron repletion at the expense of the accumulated ferritin, indicating that the ferritin-stored iron allows fast adjustment of the photosynthetic apparatus with respect to iron availability. RNAi strains that are significantly reduced in the amount of ferritin show a striking delay in the degradation of PSI under iron deficiency. Furthermore, these strains are more susceptible to photo-oxidative stress under high-light conditions. We conclude that (i) ferritin is used to buffer the iron released by degradation of the photosynthetic complexes, (ii) the physiological status of the cell determines the strategy used to overcome the impact of iron deficiency, (iii) the availability of ferritin is important for rapid degradation of PSI under iron deficiency, and (iv) ferritin plays a protective role under photo-oxidative stress conditions.     

Chlorophyll-protein composition of the thylakoid membrane from Prochlorothrix hollandica, a prokaryote containing chlorophyll b

The chlorophyll-protein complexes of the thylakoid membrane from Prochlorothrix hollandica were identified following electrophoresis under nondenaturing conditions. Five complexes, CP1-CP5, were resolved and these green bands were analyzed by spectroscopic and immunological methods. CP1 contains the photosystem I (PSI) reaction center, as this complex quenched fluorescence at room temperature, and had a 77 K fluorescence emission peak at 717 nm. CP4 contains the major chlorophyll-a-binding proteins of the photosystem II (PSII) core, because this complex contained polypeptides which cross-reacted to antibodies raised against Chlamydomonas PSII proteins 5 and 6. Furthermore, fluorescence excitation studies at 77 K indicated that only a Chl a is bound to CP4. Complexes CP2, CP3 and CP5 contained functionally bound Chl a and b as judged by absorption spectroscopy at 20 degrees C and fluorescence excitation spectra at 77 K. CP2, CP3 and CP5 all contain polypeptides of 30-33 kDa which are immunologically distinct from the LHC-II complex of higher plant thylakoids.     

Deletion Mutagenesis of the Cytochrome b559 Protein Inactivates the Reaction Center of Photosystem II

In green plant-like photosynthesis, oxygen evolution is catalyzed by a thylakoid membrane-bound protein complex, photosystem II. Cytochrome b559, a protein component of the reaction center of this complex, is absent in a genetically engineered mutant of the cyanobacterium, Synechocystis 6803 [Pakrasi, H.B., Williams, J.G.K., and Arntzen, C.J. (1988). EMBO J. 7, 325-332]. In this mutant, the genes psbE and psbF, encoding cytochrome b559, were deleted by targeted mutagenesis. Two other protein components, D1 and D2 of the photosystem II reaction center, are also absent in this mutant. However, two chlorophyll-binding proteins, CP47 and CP43, as well as a manganese-stabilizing extrinsic protein component of photosystem II are stably assembled in the thylakoids of this mutant. Thus, this deletion mutation destabilizes the reaction center of photosystem II only. The mutant also lacks a fluorescence maximum peak at 695 nm (at 77 K) even though the CP47 protein, considered to be the origin of this fluorescence peak, is present in this mutant. We propose that the fluorescence at 695 nm originates from an interaction between the reaction center of photosystem II and CP47. The deletion mutant shows the absence of variable fluorescence at room temperature, indicating that its photosystem II complex is photochemically inactive. Also, photoreduction of QA, the primary acceptor quinone in photosystem II, could not be detected in the mutant. We conclude that cytochrome b559 plays at least an essential structural role in the reaction center of photosystem II.     

THE CYANOBACTERIAL CHLOROPHYLL‐BINDING‐PROTEIN ISIA ACTS TO INCREASE THE IN VIVO EFFECTIVE ABSORPTION CROSS‐SECTION OF PSI UNDER IRON LIMITATION1

Iron availability limits primary production in >30% of the world’s oceans; hence phytoplankton have developed acclimation strategies. In particular, cyanobacteria express IsiA (iron‐stress‐induced) under iron stress, which can become the most abundant chl‐binding protein in the cell. Within iron‐limited oceanic regions with significant cyanobacterial biomass, IsiA may represent a significant fraction of the total chl. We spectroscopically measured the effective cross‐section of the photosynthetic reaction center PSI (σ_PSI) in vivo and biochemically quantified the absolute abundance of PSI, PSII, and IsiA in the model cyanobacterium Synechocystis sp. PCC 6803. We demonstrate that accumulation of IsiA results in a ～60% increase in σ_PSI, in agreement with the theoretical increase in cross‐section based on the structure of the biochemically isolated IsiA‐PSI supercomplex from cyanobacteria. Deriving a chl budget, we suggest that IsiA plays a primary role as a light‐harvesting antenna for PSI. On progressive iron‐stress in culture, IsiA continues to accumulate without a concomitant increase in σ_PSI, suggesting that there may be a secondary role for IsiA. In natural populations, the potential physiological significance of the uncoupled pool of IsiA remains to be established. However, the functional role as a PSI antenna suggests that a large fraction of IsiA‐bound chl is directly involved in photosynthetic electron transport.