Ultrastructure organization of chloroplasts in chlorotica mutants of Pisum sativum L

A study was made of chlorophyll-protein complexes of photosystems, and of ultrastructural organization of chloroplasts in pea leaves of the primary cultivar Torsdag and of its mutants, chlorotica 2004 and 2014. It has been shown that mutants accumulated 80 and 55% chlorophyll, respectively, and were able to synthesize all four types of photosystem complexes. The value of the light-harvesting antenna in mutant 2014 was close to the control one, and in mutant 2004 it increased significantly (by 30%). These changes were caused by a proportional decrease (40-50%) in any complexes in mutant 2014, whereas the number of PS-I reaction centre complexes, decreased by 50% in mutant 2004 at nearly complete storage of PS-I reaction centre complexes, decreased by 50% in mutant 2004 at nearly complete storage of PS-II complexes. The proportional decrease of PS-I and PS-II complexes in mutant chlorotica 2014 was followed by partial reduction of the entire membrane system in chloroplasts, but with a normal development of both granal and intergranal thylakoids. On the contrary, the loss of PS-I reaction centre complexes in mutant chlorotica 2004 leads to reduction of unstacked sites of thylakoids in chloroplasts. It is concluded that this effect may be associated with localization of PS-I complexes mainly in unstacked sites of thylakoids.     

Fluorescence,chlorophyll shape and number of photosystem reaction centers I and II in chlorotica mutants of Pisum sativum L

The fluorescent and absorbing properties of chloroplasts and pigment-protein complexes isolated by gel electrophoresis from pea leaves of the cultivar Torsdag and the mutants chlorotica 2004 and 2014 were studied. From the absorption and fluorescence spectra of chlorophylls and their 2nd derivatives, the range of their changes in the native state at 23 degrees C and specific maxima of fluorescence and the forms of chlorophyll of individual complexes at -196 degrees C were found. It was found that in mutant chlorotica 2004 the intensity of fluorescence of long-wave band at 745 nm (23 degrees C) and the maximum--at 728 nm (-196 degrees C) belonging to the light-harvesting complex I increased. Nevertheless, the accumulation of the chlorophyll forms in this mutant at 690, 697 and 708 nm, which make an antenna of reaction centers of photosystem (PS) I decreased. No spectral differences from the spectrum of the wild type were found in mutant chlorotica 2014, except for a weakening of interaction between the complexes of PS I and PS II. It was shown by gel electrophoresis that both mutants were capable of synthesizing any chlorophyll-protein complexes. However, the analysis of the photochemical activity of reaction centers of PS I and PS II as well as calculations of the value of the photosynthetic unit and the number of reaction centers of the photosystems enabled us to conclude that the quantity of the reaction centers of PS I in the mutant chlorotica 2004 was 1.7 times lower due to disturbance of mutations in biosynthesis or the formation of the chlorophyll a-protein complex of PS I. No primary effect of mutation of chlorotica 2014 was established. Proportional changes of all parameters in this mutant gave us the ground to consider them as secondary ones, which are caused by a decrease in chlorophyll content by half.     

The Involvement of the Complexes of Photosystems in the Organization of Chloroplast Ultrastructure in Pea Mutants

We studied the involvement of pigment-protein complexes of photosystems (PS) in the development and spatial arrangement of thylakoids in chloroplasts of pea (Pisum sativum L.) leaves. The initial line (cv. Torsdag) and its mutants, chlorotica 2004 displaying primary disturbances in the PSI reaction centers and chlorotica 2014 containing only 50% of chlorophyll and, as a sequence, the reduced amount of all pigment-protein complexes. A proportional decrease in the content of PSI and PSII complexes in the chlorotica 2014 mutant resulted in a partial reduction of the whole chloroplast membrane system, whereas grana and stroma thylakoid regions were well developed. In contrast, a loss of only 20% of chlorophyll and destruction of PSI complexes in the chlorotica 2004 mutant by 50% resulted in the destruction of stroma thylakoid regions and disturbed longitudinal thylakoid and grana orientation. It was concluded that protein-protein interactions in pigment-protein complexes played a key role in the structure of thylakoid membranes and their longitudinal orientation.     

Photosystem Damage and Spatial Architecture of Thylakoids in Chloroplasts of Pea Chlorophyll Mutants

The effect of chlorophyll–protein complexes on the ultrastructure of chloroplasts was studied in the leaves of pea, the parent cultivar Torsdag and mutants chlorotica 2004 and 2014. The mutants were shown to accumulate 80 and 55% of chlorophyll, relative to the control, while the composition of the synthesized photosystem complexes was the same as in the parent cultivar Torsdag. The size of the light-harvesting antenna was similar to the control in the 2014 mutant but considerably increased (by 30%) in the 2004 mutant. These changes were due to a proportional decrease in the number of all complexes (by 40–45%) in the 2014 mutant. At the same time, the number of reaction center complexes of photosystem I (PS I) decreased by 50% while that of photosystem II (PS II) remained virtually constant in the 2004 mutant. A proportional decrease in the number of the PS I and PS II complexes in the chlorotica 2014 mutant was accompanied by a partial reduction of the entire chloroplast membrane system against the background of normal development of both granal and intergranal sites of thylakoids. Conversely, the loss of PS I reaction centers led mainly to the reduction of the intergranal sites of thylakoids in chloroplasts. This effect is attributed to the prevalence of PS I complexes in the intergranal thylakoids.     

Spectral properties and the number of photosynthetic reaction centers in chlorophyll-deficient mutants of <Emphasis Type="Italic">Pisum sativum</Emphasis>

Spectral and photochemical properties were analyzed on intact chloroplasts and pigment-protein complexes isolated with gel electrophoresis from pea (Pisum sativum L.) leaves of parental variety Torsdag and of chlorophyll-deficient mutants chlorotica 2004 and 2014. Measurements of chlorophyll absorption and fluorescence spectra and of second derivative low-temperature (–196°C) spectra clarified exact positions of fluorescence maxima and revealed the chlorophyll forms of individual complexes in samples investigated. The chlorotica 2004 mutant, whose hybrids yield the heterosis effect, was characterized by the decreased accumulation of chlorophyll forms absorbing at 690, 697, and 708 nm, known to constitute the core antenna in the vicinity of photosystem I (PSI) reaction center. In the chlorotica 2014 mutant, whose hybrids are low productive, the interaction between PSI and PSII complexes was weakened, but no other difference from the parental variety was observed. The analysis of PSI and PSII photochemical activities, as well as estimates of light-harvesting antenna size and the number of reaction centers revealed that the chlorotica 2004 mutant is deficient in the number of PSI reaction centers by a factor of 1.7. This deficiency resulted from the mutation-induced disorder in biosynthesis of chlorophyll a-protein complex of PSI. It appears that gene interactions between the 2004 mutant and the parental variety Torsdag enhance the functional and metabolic activity of leaves in their hybrids, thereby yielding the heterosis effect.Translated from Fiziologiya Rastenii, Vol. 52, No. 2, 2005, pp. 172–183.Original Russian Text Copyright © 2005 by Ladygin, Vaishlya.This revised version was published online in April 2005 with a corrected cover date.     

Pigment–Protein Complexes and the Number of Reaction Centers of the Photosystems in Pea Chlorophyll Mutants

We studied fluorescent and absorption properties of the chloroplasts and pigment–protein complexes isolated by gel electrophoresis from the leaves of pea, the parent cultivar Torsdag and mutants chlorotica 2004 and 2014. Specific fluorescence peaks of chlorophyll forms in individual complexes have been determined from the absorption and fluorescence spectra of the chloroplast chlorophyll and their second derivatives at 23 and –196°C. The mutant chlorotica 2004 proved to have an increased intensity of a long-wave band of the light-harvesting complex I at both 23°C (745 nm) and –196°C (728 nm). At the same time, this mutant manifested a decreased accumulation of the chlorophyll forms making up the nearest-neighbor antenna of the PS I reaction center (at 690, 697, and 708 nm). No spectral differences have been revealed between chlorotica 2014 mutant and the parent cultivar. Gel electrophoresis revealed the synthesis of all chlorophyll–protein complexes in both mutants. At the same time, analysis of photochemical activity of PS I and PS II reaction centers and calculations of their number and the size of the light-harvesting antenna have shown that the number of reaction centers in the PS I of chlorotica 2004 mutant is reduced by a factor of 1.7 because its chlorophyll a–protein complex is disturbed by the mutation. The primary effect of chlorotica 2014 mutation remains unclear. The proportional changes in the content of photosystem complexes in this mutant suggest that they are secondary and result from a 50% decrease in chlorophyll content.     

Action spectra and functional antenna sizes of Photosystems I and II in relation to the thylakoid membrane organization and pigment composition

The functional organization of competent photosynthetic units in developing thylakoids from intermittent-light grown pea as well as in the unstacked, stacked and phosphorylated stacked thylakoids from its mature chloroplasts was characterized by polarographic measurements of action spectra, reaction centre contents and optical cross-sections for PS I-mediated O2 uptake and PS II-mediated O2 evolution. The minimum antenna sizes of 60 and 37 chlorophyll a molecules for PS I and PS II, respectively, were determined in developing thylakoids with a ratio of Chl a/Chl b>50. In mature chloroplasts, the embedded light-harvesting chlorophyll a/b-binding (LHC) protein complexes increased the PS I and PS II effective antenna sizes by 3–6 times depending on the thylakoid membrane organization. In unstacked thylakoids, a randomization of PS I, PS II and LHC II led to the most uniform spectral distribution of light harvesting between the two photosystems but caused the maximal difference of their antenna sizes to be 370 and 100 Chls for the competent PS I and PS II units, respectively. Following the Mg2+-induced stacking of thylakoids, opposite complementary changes of the action spectra, antenna sizes and Chl a/Chl b ratios indicated a redistribution of a LHC II pool of 100 Chl ( a + b) molecules from PS I to PS II. Unlike to the stroma-exposed PS II in unstacked thylakoids, the granal PS II units of 200 Chls demonstrated an additional 2-fold increase of the effective antenna size due to energy transfer within PS II dimers under strong background illumination, which closed >90% of reaction centres. Protein phosphorylation of the stacked thylakoids induced a significant inactivation of the O2-evolving PS II centres but did not cause complementary changes of the action spectra and antenna sizes of the competent PS I and PS II. In this case, light harvesting parameters of the O2-evolving PS II units were nearly unaffected, whereas the obvious relative increase of the PS I activity at 650 nm and its decrease at >700 nm both in the action spectrum and optical cross-section measurements might suggest a substitution of PS I units in the O2-reducing fraction by another distinct fraction of -type which in turn is not the same to PS I units in unstacked thylakoids.     

Pigment accumulation and functional activity of chloroplasts in common Pisum sativum L. mutants with low chlorophyll level (chlorotica)

Pea mutants chlorotica 2004 and 2014 with a low content of chlorophyll were studied. The mutant 2004 has light green leaves and stem, and the mutant 2014 has yellow green leaves and stem. They accumulate approximately 80 and 50% chlorophylls of the parent form of pea Torsdag cv. The content of carotene in carotenoids of the mutant 2004 was much lower, and the accumulation of lutein and violaxanthine was increased. The accumulation of all carotenoids in the mutant 2014 decreased almost proportionally to a decrease in the chlorophyll content. The rate of CO2 evolution in mutant chlorotica 2004 and 2014 was established to be lower. The quantum efficiency of photosynthesis in the mutants was 29-30% lower as compared to the control, and in hybrid plants it was 1.5-2-fold higher. It is assumed that the increase in the activity of the night-time respiration in gas exchange of chlorotica mutants and the drop of photosynthesis lead to a decrease in biomass increment. The results obtained allow us to conclude that the mutation of chlorotica 2004 and 2014 affects the genes controlling the formation and functioning of different components of the photosynthetic apparatus.     

Changes in the Biochemical Composition, Structure, and Function of Pea Leaf Chloroplasts in Iron Deficiency and Root Anoxia

A combined effect of iron deficiency and root anoxia on the biochemical composition, function, and structure of pea leaf chloroplasts was studied. It was found that the chlorosis of apical leaves in response to iron deficiency was determined by the reduction of light-harvesting complexes I and II. Under root anoxia, complexes of the reaction centers of photosystems I and II degraded first. Weak activity was preserved even in yellow and white leaves under the effect of both factors. The ultrastructure of leaf chloroplasts gradually degraded. Initially, intergranal thylakoid sites were reduced, and the longitudinal orientation of grana was disturbed. However, yellow and white leaves still retained small thylakoids and grana. It is concluded that the degrading effects of iron deficiency and root anoxia on the complex composition and leaf chloroplast structure and function are additive because of their autonomous mechanisms.     

Changes in the biochemical composition, structure, and function of pea leaf chloroplasts in iron deficiency and root anoxia

A combined effect of iron deficiency and root anoxia on the biochemical composition, function, and structure of pea leaf chloroplasts was studied. It was found that the chlorosis of apical leaves in response to iron deficiency was determined by the reduction of light-harvesting complexes I and II. Under root anoxia, complexes of the reaction centers of photosystems I and II degraded first. Weak activity was preserved even in yellow and white leaves under the effect of both factors. The ultrastructure of leaf chloroplasts gradually degraded. Initially, intergranal thylakoid sites were reduced, and the longitudinal orientation of grana was disturbed. However, yellow and white leaves still retained small thylakoids and grana. It is concluded that the degrading effects of iron deficiency and root anoxia on the complex composition and leaf chloroplast structure and function are additive because of their autonomous mechanisms.     

Structural and functional organization of chloroplasts in leaves of Pisum sativum L. under conditions of root hypoxia and iron deficiency

A combined effect of iron deficiency and root hypoxia on the biochemical composition activity and structure of chloroplasts in pea leaves have been studied. Both factors are shown to affect the accumulation of chlorophyll causing leaf chlorosis. At iron deficiency chlorosis occurs from the top of plant leaves. At root hypoxia chlorosis starts from the lower strata. At a combined action of both factors the destructive effects are summarized. It was established that light-harvesting complexes of photosystems were reduced stronger at iron deficiency, while complexes of reaction centers of photosystem I and photosystem II are lessened at root hypoxia. Nevertheless, even at a combined effect of both factors yellow leaves preserved small amounts of any pigment-protein complexes and their functional activities. The ultrastructure of chloroplasts during leaf chlorosis was gradually reduced. At first, intergranal sites of thylakoids and then granal ones were destroyed, that was typical of iron deficiency. However, even yellow and almost white leaves kept small thylakoids, capable of forming stacking and small grana made of 2-3 thylakoids. It has been concluded that the destructive effects are summarized due to different kinds of action of iron deficiency and root hypoxia on the structure and functioning of leaves at their combined action.     

Functional effects of chemical modification of unstacked and stacked chloroplasts with glutaraldehyde.

Although glutaraldehyde alkylates protein NH₂ groups to the same extent in unstacked and stacked thylakoids, the photosynthetic electron transport of the stacked membranes is always more inhibited. Inhibition of photosystem II electron transport, measured in the presence of lipophilic Hill oxidants, is 20–30% in unstacked and 60–70% in stacked thylakoids. Photosystem I electron transport is nearly completely inhibited in both preparations, but in the case of stacked thylakoids maximal inhibition occurs at a lower glutaraldehyde level than in unstacked thylakoids. In contrast, the photooxidation of the reaction center chromophore of photosystem I (P700) is unaffected by the glutaraldehyde treatment of either stacked or unstacked chloroplasts. The results are discussed with regard to the accessibility of membrane sites to exogenous electron transport cofactors, in view of the observation that N-methylphenazonium methosulfate, a quencher of electronically excited chlorophyll a, partitions more easily into the pigment domains of the glutaraldehyde-fixed unstacked thylakoids.     

Functional assembly in vitro of phycobilisomes with isolated photosystem II particles of eukaryotic chloroplasts

Phycobiliproteins obtained by dissociation of phycobilisomes were reassociated in vitro with intact thylakoids or isolated photosystems I and II preparations obtained from cyanophytes (prokaryotes) or green algae (eukaryotes) to form bound phycobilisome complexes. Energy transfer from Fremyella diplosiphon phycobiliproteins to chlorophyll a of reaction centers I and II was measured in: complexes containing intact thylakoids of the cyanophytes F. diplosiphon or Anacystis nidulans and the eukaryotic algae Euglena gracilis and mutants of Chlamydomonas reinhardtii; complexes containing isolated photosystem II particles of A. nidulans or C. reinhardtii; and complexes containing reaction center I of F. diplosiphon or C. reinhardtii. Energy transfer from phycoerythrin to chlorophyll a of photosystem II could be demonstrated in complexes containing phycobilisomes bound to cyanophyte thylakoids or isolated photosystem II particles of A. nidulans or C. reinhardtii. Bound phycobilisomes did not transfer energy to photosystem II within green algae thylakoids containing altered forms of light-harvesting chlorophyll a/b-protein complex (LHC) II antenna, reduced amounts of LHC II, or chlorophyll b, or chlorophyll b-less mutants, nor to chlorophyll a of photosystem I of intact thylakoids or isolated reaction centers. We conclude that phycobilisomes can form a specific and functional association with photosystem II particles of both cyanophytes and eukaryotic thylakoids. This interaction appears to be hindered by the presence of LHC II antenna in the eukaryotic thylakoids.     

The Effect of Root Hypoxia and Iron Deficiency on the Photosynthesis, Biochemical Composition, and Structure of Pea Chloroplasts

The combined effect of root hypoxia and iron deficiency on biochemical composition, photosynthetic indices, and structure of pea (Pisum sativum L.) chloroplasts were investigated. Both factors suppressed chlorophyll accumulation and leaf photosynthetic activity, causing chlorosis. It was shown, that iron deficiency reduced more severe the light-harvesting complexes of photosystems (PS), and root hypoxia, the reaction center complexes of the photosystem I (PSI) and photosystem II (PSII). The combined action of both factors was stronger than the effect of each factor. However, even in yellow and almost white leaves, chloroplasts contained small amounts of all pigment–protein complexes and maintained weak photosynthetic activity, although their structure was poorly developed and comprised only vesicles and small thylakoids capable to form contacts and small grana. The conclusion is that the mechanisms of root hypoxia and iron deficiency destructive action are different and these factors differently and independently influenced leaf chloroplasts.     

Structural-functional organization of chloroplasts in leaves of xantha-702 mutant of Gossypium hirsutum L

For cotton mutant xantha (Gossypium hirsutum L.), it has been established that synthesis of 5-aminolevulinic acid was blocked in the light. In the light this mutant accumulates chlorophyll by 30 times lower as compared to the parent type. In mutant xantha, a very few pigment-protein complexes of PS-I and PS-II are formed in chloroplasts, and formation of membrane system in these is blocked at the early stages, in most cases, at the stage of bubbles and single short thylakoids. Functional activity of reaction centers of PS-I and PS-II is close to zero. Only light-harvesting chlorophyll-a/b protein complexes of the two photosystems are formed in mutant xantha plastid membranes with maximum chlorophyll fluorescence at 728 and 681 nm, respectively. It has been concluded that in mutant xantha genetic block of 5-aminolevulinic acid biosynthesis in the light disturbs the formation and functioning of the complexes of reaction centers of PS-I and PS-II, hindering the development of the whole membrane system in chloroplasts, causing a sharp decrease in productivity.     

Spectral characteristics and the structure of chloroplasts upon blocking the early stages of chlorophyll biosynthesis

The cotton mutant xantha (Gossypium hirsutum L.) with the blocked synthesis of 5-aminolevulinic acid in the light has been shown to accumulate chlorophyll 30 times less than the parent type. In chloroplasts of the mutant xantha, the formation of the membrane system is blocked at the earliest stages, mainly at the stage of bubbles and single short thylakoids. Only light-harvesting chlorophyll-a/b-protein complexes I and II with chlorophyll fluorescence maxima at 728 and 681 nm, respectively, are formed in plastid membranes of the mutant. It has been concluded that the genetic block of chlorophyll biosynthesis in the mutant xantha disturbs the formation and functioning of the complexes in reaction centers of PS-I and PS-II, inhibiting the development of the whole membrane system of chloroplasts at the stage of bubbles and single thylakoids.     

Functional organization of thylakoid membranes in viable pea mutants with low chlorophyll content

The functional organizations of thylakoid membranes from wild type pea ( Pisum sativum L. cv. Kapital) and two viable mutants with low chlorophyll (Chl) contents were compared. Nuclear mutations in mutants 7 and 42 led to two- and three-fold decrease in total chlorophyll content, respectively. In spite of low Chl content mutants showed 80% photosynthetic activity, biological productivity, and seed production. It has been shown that mutant membranes differed from that of wild type by Chl distribution between the pigment-protein complexes and by stoichiometry of the main electrontransport complexes. The ratio photosystem I (PSI): photosystem II (PSII): cytochrome (Cyt) bjf complex: Chl was 1:1.1:1.2:650 in wild type chloroplasts, 1:1.8:1.7:600 in mutant 7 , and 1:1.5:1.9:350 in mutant 42 . PSI- and PSII-dependent electron-transport activities were enhanced in the mutants per mg Chl in proportion to number of reaction centers. The activity of the non-cyclic electron-transport chain increased in proportion to PSII and Cyt bjf complexes. The amount of ATP synthetase per unit of Chl as estimated by H⁻ATPase activity was much greater in mutant thylakoids, which is favorable for photosynthetic energy transduction. The low content of the light-harvesting complexes (LHC) in mutants is compensated by an increase of the number of PSII and Cyt bjf complexes, which eliminates the bottleneck at the site of plastoquinone oxidation.     

The coleoptile chloroplast: Distinct distribution of xanthophyll cycle pigments,and enrichment in Photosystem II

Recent studies have shown that coleoptile chloroplasts operate the xanthophyll cycle, and that their zeaxanthin concentration co-varies with their sensitivity to blue light. The present study characterized the distribution of photosynthetic pigments in thylakoid pigment–protein complexes from dark-adapted and light-treated coleoptile and mesophyll chloroplasts, the low temperature fluorescence emission spectra, and the rates of PS I and PS II electron transport in both types of chloroplasts from 5-day-old corn seedlings. Pigments were extracted from isolated PS I holocomplex, LHC IIb trimeric and LHC II monomeric complexes and analyzed by HPLC. Chlorophyll distribution in coleoptile thylakoids showed 31% of the total collected Chl in PS I and 65% in the light harvesting complexes of PS II. In mesophyll thylakoids, the values were 44% and 54%, respectively. Mesophyll and coleoptile PS I holocomplexes differed in their Chl t a/Chl t b ratios (8.1 and 6.1, respectively) and -carotene content. In contrast, mesophyll and coleoptile LHC IIb trimers and LHC II monomers had similar Chl t a/Chl t b ratios and -carotene content. The three analyzed pigment–protein complexes from dark-adapted coleoptile chloroplasts contained zeaxanthin, whereas there was no detectable zeaxanthin in the complexes from dark-adapted mesophyll chloroplasts. In both chloroplast types, zeaxanthin and antheraxanthin increased markedly in the three pigment–protein complexes upon illumination, while violaxanthin decreased. In mesophyll thylakoids, zeaxanthin distribution as a percentage of the xanthophyll cycle pool was: LHC II monomers > LHC IIb trimers > PS I holocomplex, and in coleoptile thylakoids, it was: LHC IIb trimers > LHC II monomers = PS I holocomplex. Low temperature (77 K) fluorescence emission spectra showed that the 686 nm emission of coleoptile chloroplasts was approximately 50% larger than that of mesophyll chloroplasts when normalized at 734 nm. The pigment and fluorescence analysis data suggest that there is relatively more PS II per PS I and more LHC I per CC I in coleoptile chloroplasts than in mesophyll chloroplasts. Measurements of t in vitro uncoupled photosynthetic electron transport showed approximately 60% higher rates of electron flow through PS II in coleoptile chloroplasts than in mesophyll chloroplasts. Electron transport rates through PS I were similar in both chloroplast types. Thus, when compared to mesophyll chloroplasts, coleoptile chloroplasts have a distinct PS I pigment composition, a distinct chlorophyll distribution between PS I and PS II, a distinct zeaxanthin percentage distribution among thylakoid pigment–protein complexes, a higher PS II-related fluorescence emission, and higher PS II electron transport capacity. These characteristics may be associated with a sensory transducing role of coleoptile chloroplasts.     

Properties of thylakoids and thylakoid particles derived from structurally different chloroplasts

Structurally and functionally different tobacco chloroplasts were subjected to digitonin treatment and subsequent fractional centrifugation. The light-harvesting $\text{chlorophyll}\text{a}\text{chlorophyll}\text{b-}\text{protein}$ complex was found to be enriched in the most dense fraction regardless of the presence of grana in the original preparation. It is suggested that isolated thylakoid membranes and fragments thereof which contain sufficient light-harvesting protein may, under appropriate ionic conditions, form aggregates even when they originate from unstacked thylakoid systems. Comparative studies of fluorescence properties and polypeptide composition of the thylakoids suggest that the light-harvesting protein does not contribute significantly to the fluorescence spectrum of isolated chloroplasts as long as this protein is intimately associated with the Photosystem II (PS II) pigment-protein complex responsible for the 685 nm emission. While the PS II-deficient mutant chloroplasts of the variegated tobacco variety NC 95 lacked both the 685 nm fluorescence component and two or three PS II proteins, one of these proteins was found to be very prominent in our chlorophyll b-deficient mutant thylakoids which also displayed an intense 685 nm fluorescence peak. This correlation supports the contention that a 45 kdalton polypeptide is an apoprotein of pigments associated with the PS II reaction center.     

Protein carriers of the electron transport chain of non-photosynthesizing mutants of Chlamydomonas reinhardii]

The amounts of ferredoxin, plastocyanin, ferredoxin-NADP-reductase were determined by electrophoresis and differential spectroscopy. The cytochrome levels in the chloroplasts of non-photosynthesizing mutants Chlamydomonas reinhardii were determined both in active and inactive photosystems. It was shown that the loss of PS-1 activity did not affect the amount and activity of the electron carriers. The disturbances of the donor side of PS-2 in the mutants were accompanied by a loss of the reaction center activity and by a decrease of cytochrome b599. The amounts of other protein components in the mutants with inactive PS-2 remained unchanged. The disturbances in the cytochrome c553 content presumably blocked the electron transfer between the photosystems but did not affect the activity of the reaction centers of PS and the levels of other carriers of the chloroplast electron transport chain.