Conformational studies of sequential polypeptides containing lysine and tyrosine

The conformation of three sequential copolypeptides, poly(L -tyrosyl-L -lysine), poly(L -tyrosyl-L -lysyl-L -lysine), and poly[L -tyrosyl-(L -lysyl)₂-L -lysine] have been studied by a variety of techniques, including CD, ir spectroscopy, analytical ultracentrifugation, and x-ray diffraction. Depending upon the pH and sovent composition, poly(L -tyrosyl-L lysyl-L -lysine) and poly [L -tyrosyl-(L lysyl)₂-L -lysine] can adopt either the α-helical or random-coil conformation, while poly(L -tyrosyl-L -lysine) forms either inter- or intramolecular β-structures.     

Block oligopeptides (L-lysyl)m-(L-alanyl)n-L-Tyrosyl-(L-Alanyl)n-(L-Lysyl)m. II. Circular dichroism and pulsefluorimetry conformational studies

The conformation of chromatographically pure block oligopeptides (L -lysyl)_m-(L -alanyl)_n- L -tyrosyl-(L -alanyl)_n-(L -lysyl)_m with n = 3 and m = 6 or 3 is investigated. By circular dichroism it is shown that these peptides may exhibit a partially α-helical structure depending upon pH, ionic strength, solvent, and temprerature. An attempt is made to describe the helical content of these small peptides by utilizing the data obtained on high-molecular-weight poly(L -lysine). By measurement of the quantum yield and the decays of the peptides fluorescence, it is shown that, in aqueous solution, at neutral pH, the fluorescence of the peptides is quenched by interactions with the peptide carbonyl groups. The decays are multiexponential, which shows the presence of several conformations of the phenolic chromophore relative to the peptide chain. The addition of methanol, which induced the helix formation, decreases the quenching of the fluorescence and the multiexponential character of the decays. In presence of sodium hydroxide, which further increases the helical content of the peptides, a dynamic quenching occured that can be attributed to interactions between the phenol hydroxyl group of tyrosine (ith residue) and the ε-amino groups of the (i+4)th and (i -4)th lysyl residues.     

The beta conformation of polypeptides of valine, isoleucine, and threonine in solution and solid-state: optical and infrared studies.

Water-soluble polypeptides of L -valyl and L -isoleucyl residues flanked with DL -lysyl blocks [poly(DL Lys · HCl)_x–poly(L Val)_y–poly(DL Lys · HCl)_x, poly(DL Lys · HCl)_x–poly-(L Ile)_y–poly(DL Lys · HCl)_x] and homopoly(L -threonine) were prepared. The β conformation of these polymers in water, as well as in aqueous methanol, was confirmed by infrared spectroscopy and circular dichroism studies. The optical properties of these valyl and isoleucyl polypeptides were quite different from those of previously reported synthetic homopolypeptides in the β structure. Their differences could be explained by the presence of a “single extended β chain” without either intra- or interchain association.     

Block oligopeptides (L-lysyl)m-(L-alanyl) L-tyrosyl-(L-alanyl)n- (L-lysyl)m. I. Synthesis through fragment condensation and characterization

Model peptides containing one aromatic residue were synthesized and characterized in order to investigate their interactions with polynucleotides. Chromatographically pure block oligopeptides (L -alysyl)_m-(L -alanyl)_n- L -tyrosyl- (L -alanyl)_n, with n = 3 and m=3 or 6, were prepared by fragments condensation using the mixed anhydride method. The protected fragments were prepared by stepwise addition of amino acid residues through the dicyclohexylcarbodiimide method. The purity of the intermediate coupling product was analyzed by gradient elution chromotography on carboxylmethylcellulose. Both block oligopeptides were isolated by preparative chromatography on carboxymethylcellulose. The different features of these syntheses are discussed.     

Synthesis and conformational study of sequential polypeptides, (L-ala-L-val-gly)n and (L-val-L-ala-gly)n.

As an approach for elucidating the role of sequences of amino acids in protein structures, model polypeptides having the same composition but different sequences of amino acids, (L -Ala-L -Val-Gly)_n and (L -Val-L -Ala-Gly)_n, have been prepared by the method involving facile monomer synthesis using N-carboxy α-amino acid anhydrides and N-hydroxysuccinimide esters. The yields and the molecular weights of the polypeptides formed by polycondensation do not depend on the monomer concentrations, but on the sequences of the amino acids in the monomers. Infrared spectra in the solid state showed that (L -Ala-L -Val-Gly)_n can take the α-helical conformation but (L -Val-L -Ala-Gly)_n cannot. The results suggest that the conformations of polypeptides are influenced by the sequences of the amino acids in the polypeptides.     

Carrier design: Conformational studies of amino acid (X) and oligopeptide (X-DL-Alam) substituted poly(L-lysine)

The present study was undertaken to examine the influence of the reversal of the sidechain sequential order on the conformation of branched polypeptides. At the same time, the influence of the optically active amino acid joined directly to the poly (L -Lys) backbone and the DL -Ala oligomer grafted as chain-terminating fragment were separately analyzed. Therefore two sets of polypeptides were synthesized corresponding to the general formula poly [Lys-(X_i,)] (XK) and poly[Lys-(DL -Ala_m-X_i)] (AXK) when X = Ala, D -Ala, Leu, D -Leu, Phe, D -Phe, Ile, Pro, Glu.,D -Glu, or His. For coupling amino acid X to polylysine, three types of active ester methods were compared: the use of pentafluorophenyl or pentachlorophenyl ester, and the effect of the addition of an equimolar amount of 1-hydroxybenzotriazole. After cleavage of protecting groups, AXK polypeptides were synthesized by grafting short oligo (DL -Ala) chains to XK by using N-carboxy-DL -Ala anhydride. The CD measurements performed in water solutions of various pH values and ionic strengths were used for classification of the polypeptide conformations as either ordered (helical) or unordered. Different from what was observed with the unsubstituted poly (L -Lys), poly[Lys-(X_i)] type polypeptides can adopt ordered structure even under nearly physiological conditions (pH 7.3, 0.2M NaCl). These data suggest that the introduction of amino acid residue with either (ar) alkyl side chain (Ala, Leu, Phe) or negatively charged side chain (Glu) promotes markedly the formation of ordered structure. Comparison of chiroptical properties of poly [Lys- (DL -Ala_m-X_i)] and of poly [Lys- (X_i)] reveals that side-chain interactions play an important role in the stabilization of ordered solution conformation of AXK type branched polypeptides. The results give rather conclusive evidence that not only hydrophobic interactions, but also ionic attraction, can be involved in the formation and stabilization of helical conformation of branched polypeptides. © 1993 John Wiley & Sons, Inc.     

Gibberellin-photoaffinity labelling of two polypeptides in plant plasma membranes

Two polypeptides of M _r 68 kDa and 18 kDa were gibberellin (GA)-photoaffinity labelled in vitro in plasma membrane preparations from oat ( Avena sativa L.) aleurone and from leaves and stems of wild-type and GA-sensitivity mutants of different species. Labelling of these polypeptides could be competed by biologically active, but not by inactive, GAs, indicating the likely biological significance of these interactions. On 2-dimensional gels the radiolabelled polypeptides were each resolved as one intensely labelled low abundance spot with a slightly lower pI form adjacent to it. There was a strong pH dependency for both labelling events, which correlated well with pH values at which GA are known to be most biologically active. A semi-dwarf GA-sensitivity mutant of sweet pea ( Lathyrus odoratus L.), lb , showed reduced photoaffinity labelling of both polypeptides compared with the wild type, Lb . In the GA-insensitive Arabidopsis thaliana mutant, gai , the level of labelling was the same as in wild type, GAI . This is the first report of GA-binding proteins in plant plasma membranes. Some preliminary sequence data are given for one of the labelled polypeptides. We discuss these mutants and consider their possible roles in GA perception or action.     

Expression of nuclear and chloroplastic genes coding for fraction-1 protein in somatic hybrids of Nicotiana tabacum + rustica

In the sexual interspecific cross, Nicotiana rustica L.xN. tabacum L., N. rustica can serve as the female but not as the male parent. By fusion of protoplasts, the barrier to fertilization was overcome and somatic hybrids containing N. tabacum cytoplasm were produced as shown by isoelectric focusing of the Fraction-1 protein (F-1-protein). All somatic hybrids displayed polypeptides of the large subunit of F-1 protein (which is coded by the chloroplast genome) characteristic of only one or the other parental species. Two hybrids had large subunits of the N. tabacum type and two hybrids had those of the N. rustica type. Three hybrids contained three smallsubunit polypeptides (coded by the nuclear genome), one being characteristic of N. rustica, one characteristic of N. tabacum, and one with an isoelectric point common to both species. A fourth hybrid contained only two small-subunit polypeptides of the N. tabacum type but in a F-1 protein macromolecule whose large subunits were of the N. rustica type. One somatic hybrid was self-fertile and its F₂ progeny contained large- and small-subunit polypeptides indistinguishable in their isoelectric points from those in the parent F₁ hybrid. All somatic hybrids showed an aneuploid chromosome number and morphological characteristics intermediate between those of N. rustica and N. tabacum.     

Synthesis of the vasoactive intestinal peptide (VIP) : IV. The sequence 14–28

A protected pentadecapeptide with the C-terminal sequence of the vasoactive intestinal peptide (VIP) was prepared by coupling the tetrapeptide derivative t-butyloxycarbonyl- -arginyl-N-benzyloxycarbonyl- -lysyl- -glutaminyl- -methionine azide to the partially deprotected hendecapeptide -alanyl- -valyl-N-benzyloxycarbonyl- -lysyl-N-benzyloxycarbonyl- -lysyl- -tyrosyl- -leucyl- - asparaginyl- -seryl- -isoleucyl- -leucyl- -asparaginamide. The preparation of the protected tetradecapeptide t-butyloxycarbonyl-N-benzyloxycarbonyl- -lysyl- -glutaminyl- -methionyl- -alanyl- -valyl-N- benzyloxycarbonyl- -lysyl-N-benzyloxycarbonyl- -lysyl- -tyrosyl- -leucyl- -asparaginyl- -seryl- -isoleucyl- -leucyl- -asparaginamide is also reported. The protecting groups were removed from samples of the tetradeca- and pentadecapeptides. The resulting free peptides showed, although at high dose levels, increase of visceral blood flow and reduction of blood pressure in the dog, and also relaxation of different smooth muscle preparations, which are the characteristic biological activities of VIP.     

Block sequential polypeptides of L-alanine and glycine with D, L-glutamic acid

Sequential polypeptides of L -alanine(A) and glycine(G), which were incorporated between two blocks of poly(D ,L -glutamic acid) (DL), were synthesized by applying Merri-field's solid-phase method. On the basis of optical rotatory dispersion criteria, DL(A)₃₈-DL was found to assume the α-helix in the whole range of the water-methanol system; whereas other block sequential polypeptides were found to assume the random-coiled conformation in water and partly the α-helix at the high methanol content. The stability of the α-helix decreased in the order: DL(A)₃₈DL, DL(A₂G)₁₀DL, DL(A₂G)₆DL, and DL(A₃G)₇DL. This phenomenon may be explained in terms of the dependence of hydrophobic bonding between the C₃H group of the ith L -alanine regularly arranged on the surface of the α-helix and the C₂H group of the (i + 3)th residue on whether the residue is alanine or glycine. The role which the methanol plays in stabilizing the α-helix is also discussed.     

Protein profiling in F<Subscript>1</Subscript> and F<Subscript>2</Subscript> generations of two tomato genotypes differing in ripening time

Pericarp polypeptide profiles were analyzed at three ripening stages in the F₁ hybrid and the F₂ population from the cross between the accessions: LA1385 (Lycopersicon esculentum var. cerasiforme) and 804627 (L. esculentum, a homozygous genotype for the nor mutant). Six polymorphic polypeptides were observed in LA1385, while no polymorphic polypeptides among ripening stages was observed in 804627. On the other hand, some polypeptides in the F₁ hybrid were not observed in the parents whereas others were present in both parental genotypes and were unnoticeable in the hybrid genotype. From a cluster analysis on the protein profiles of the F₂ population, the differential expression of proteins allowed to distinguish mature green (MG) stage from the others two stages, while for breaker stage (BR) and red ripe stage, the genetic background was more important in forming groups. The differential expression of proteins could be associated with fruit morphology traits such as a 72 kDa polypeptide present in MG stage with fruit diameter, height and mass and a 47 kDa polypeptide found in BR with fruit shelf life.     

A high-resolution 13C-nmr study of collagenlike polypeptides and collagen fibrils in solid state studied by the cross-polarization–magic angle-spinning method. Manifestation of conformation-dependent 13C chemical shifts and application to conformational characterization

We have recorded high-resolution ¹³C-nmr spectra of collagen fibrils in the solid state by the cross-polarization–magic-angle-spinning(CP–MAS)method and analyzed the spectra with reference to those of collagenlike polypeptides. We used two kinds of model polypeptides to obtain reference ¹³C chemical shifts of major amino acid residues of collagen (Gly, Pro, Ala, and Hyp): the 3₁-helical polypeptides [(Gly)_nII, (Pro)_nII, (Hyp)_n, and (Ala? Gly? Gly)_nII], and the triple-helical polypeptides [(Pro? Gly? Pro)_n and (Pro? Ala? Gly)_n]. Examination of the ¹³C chemical shifts of these polypeptides, together with our previous data, showed that the ¹³C chemical shifts of individual amino acid residues are the same, within experimental error (±0.5 ppm), among different polypeptides with different primary sequences, if the conformations are the same. We found that the ¹³C chemical shifts of Ala residues of the 3₁-helical (Ala? Gly? Gly)_n and triple-helical (Pro? Ala? Gly)_n are significantly displaced, compared with those of the α-helix, β-sheet, and silk I form, and can be utilized as excellent probes to examine conformational features of collagen-like polypeptides. Further, the ¹³C chemical shifts of Gly and Pro residues in the triple-helical polypeptides are substantially displaced from those found in (Gly)_nII and (Pro)_nII of the 3₁-helix, reflecting further conformational change from the 3₁-helix to the supercoiled triple helix. In particular, the ¹³C chemical shifts of Gly C ? O carbons of the triple-helical polypeptides are substantially displaced upfield (4.1–5.1 ppm), with respect to those of the 3₁-helical polypeptides. These displacements are interpreted by that Gly C ? O of the former is not involved in NH …? O ? C hydrogen bonds, while this carbon of the latter is linked by these kinds of hydrogen bonds. On the basis of these ¹³C chemical shifts, as reference data for the collagenlike structure, we were able to assign the ¹³C-nmr peaks of Gly, Ala, Pro, and Hyp residues of collagen fibrils, which are in good agreement with the values expected from the model polypeptides mentioned above. We also discuss a plausible conformational change of collagen fibrils during denaturation.     

Purification and properties of two truncated endoglucanases produced in Escherichia coli harbouring Clostridium cellulolyticum endoglucanase gene celCCD

The endoglucanase gene, celCCD, of Clostridium cellulolyticum has been expressed in Escherichia coli. Multiple active polypeptides were detected in the E. coli cells. The relative molecular mass (M_r) of two major active polypeptides were 56 000 (D56) and 38 000 (D38), which were smaller than the deduced M_r of the mature protein (63 401). D56 and D38 were purified from the periplasmic fraction. The N-terminal sequences of the two purified polypeptides were identical to that of the mature endoglucanase (Ala-Ile-Asn-Ser-Gln-Asp-Met-Val---) deduced from the nucleotide sequence. These data indicated that these polypeptides were produced by processing the original mature protein in the C-terminal region. The enzymatic properties of these two polypeptides were very similar, except that the specific activity of D38 was 2–3.5-fold higher than that of D56, and D38 was more heat stable than D56. Correspondence to: T. Kodama     

Evolution of fraction 1 protein in the genus Lycopersicon

The large- and small-subunit polypeptide composition of fraction 1 protein contained in seven species of Lycopersicon and Solanum pennellii was determined by electrofocusing. The eight species of protein had large subunits composed of three polypeptides separated by about 0.05 pH unit, but there was no difference in the isoelectric points of the clusters of three polypeptides. By this criterion, no surviving mutations have appeared in the extranuclear DNA coding for the cluster of large-subunit polypeptides during a period of evolution which generated the eight species of plants. The genus Lycopersicon appears to be much younger than its sister genus Nicotiana in the family Solanaceae, where four types of polypeptide clusters have evolved. Three different small-subunit polypeptides whose isoelectric points are coded by nuclear DNA have arisen among the seven Lycopersicon species, and L. hirsutum and S. pennellii have proteins containing single polypeptides and are therefore considered older than L. chilense, L. chimielewskii, and L. parviflorum, whose proteins contain two polypeptides. L. cheesemanii, L. pimpinellifolium, and L. esculentum (and probably L. peruvianum) seem to be the most recently evolved species since their fraction 1 proteins have small subunits composed of three polypeptides.This research was supported by NSF Grant 75-07368 and Contract No. EY-76-S-03-0034, P. A. #8, from the Department of Energy.     

Polypeptide composition and biosynthesis of the yolk proteins in the firebrat,Thermobia domestica (insecta,thysanura)

Ion-exchange chromatography of crude ovarian extracts of the primitive insect Thermobia domestica allowed the separation, in native conditions, of major and minor vitellins of molecular weights of 300,000 and 430,000, respectively. Their polypeptide subunits were determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and immunotransfer using an antiserum prepared against major vitellin. This protein was resolved into large (M_r 166,000–212,000) and small (around M_r 50,000) polypeptides. Minor vitellin, on the other hand, exclusively contained small polypeptides that are immunologically different from those of the major vitellin. Vitellogenin polypeptides from the hemolymph of mature females exhibited electrophoretic mobilities and immunological properties similar to vitellin polypeptides. Pulse-chase experiments showed that the female fat body synthesizes radioactive and immunoprecipitable proteins, whose polypeptide pattern is close to that of the major vitellogenin. However, part of the primary vitellogenic polypeptides, at M_r 210,000 and 212,000, is rapidly processed to M_r 176,000 and 182,000 subunits. These two polypeptides, as well as the precursors, enter into the composition of the major hemolymph vitellogenin. Finally, processing of the still uncleaved 210,000–212,000 polypeptides takes place in the ovary, which performs the same step of vitellogenin maturation as the fat body.     

Variation in mitochondrial translation products in fertile and cytoplasmic male-sterile sugar beets

Summary Intact and functional mitochondria were isolated from sugar beet plants (Beta vulgaris L.) containing normal fertile (F) or cytoplasmic male-sterile (S₁–S₄) cytoplasms. Incorporation of ³⁵S-methionine by mitochondria isolated from both roots and leaves showed approximately 20 major and ten minor translation products. Comparison of the polypeptide synthesis patterns produced by leaf mitochondria from fertile plants of three different species within the genus Beta revealed several taxonomically related differences. Contrary to this, the patterns of polypeptides synthesized by mitochondria from roots and leaves of sugar beet plants containing the F and S₁–S₄ cytoplasms were very similar; in the S₁ and S₂ cytoplasms no qualitative, and only a few quantitative, differences from the F cytoplasm were observed. Thus, in these cases, cytoplasmic male sterility in sugar beet is not correlated with the constitutive expression of variant polypeptides. In the S₃ cytoplasm, however, an additional 6 kDa polypeptide was synthesized and in the S₄ cytoplasm an additional 10 kDa polypeptide was observed when compared with the F cytoplasm. The expression of cytoplasmic male sterility in sugar beet may be associated with these variant polypeptides. The mitochondrial polypeptides synthesized were identical in plants with different nuclear backgrounds but with identical S₁ cytoplasms. Mitochondria from plants with variants of the S₄ cytoplasm in the same nuclear genotype also showed identical patterns of polypeptide synthesis, including the synthesis of the 10 kDa S₄-specific polypeptide. Pulse-chase experiments did not affect the synthesis of this polypeptide.     

Btl, a structural gene for the major 39–44 kDa amyloplast membrane polypeptides

The aim of the present work was to investigate the relationship between the Btl gene (Btl) and the major 39–44 kDa amyloplast membrane polypeptides which were deficient in amyloplast membranes of brittlel (btl) kernels of maize (Zea mays L.). A rapid yet gentle procedure for the isolation of amyloplasts from immature kernels is described. These amyloplasts were relatively free of contamination by other cellular components, and immunological studies showed that they contained polypeptides which reacted with antibodies to maize starch branching enzyme and ADP-Gle pyrophosphorylase. Purified membranes isolated from the amyloplast contained a poly-peptide which reacted with antibodies to the P_i-translocator from spinach chloroplasts. However, a cluster of 39–44 kDa polypeptides accounted for about 40% of the total amyloplast membrane protein from W64A kernels. These polypeptides were specifically recognized by antibodies raised against a fusion protein consisting of 56 amino acids of the carboxyl terminus of the BTI protein and glutathione S-transferase. The BT1 antibodies also reacted with the abundant polypeptides in amyloplast membranes from hybrid kernels (Doebler 66XP and Pioneer 3780), and the shrunkenl and shrunken2 mutant genotypes, but no BTl reacting polypeptides were present in amyloplast membranes from btl mutant kernels. We were unable to detect BTl by the immunoblot procedure in microsomal membranes from embryo and pericarp tissues from the kernel, from seedling roots and shoots, or in membranes from mitochondria and chloroplasts. The same BTl immunoblot pattern was obtained for proteins extracted from microsomal membranes from developing endosperm and from purified amyloplast membranes. A linear relationship between the number of copies of Btl alleles and the levels of BTl in endosperm microsomal membranes was demonstrated in a gene dosage series. BTl was not extracted from amyloplast membranes by chloroform/methanol or by alkaline buffer at pH 11.5, but was partially extracted by 0.1 M NaOH. These lines of evidence support the conclusion that Btl is the structural gene for the major 39–44 kDa amyloplast membrane polypeptides and that these polypeptides are integral proteins specific to amyloplast membranes from the endosperm.     

Unassembled polypeptides of the plastidic ribosomes in heat-treated 70S-ribosome-deficient rye leaves

The polypeptides of the subunits of 70S ribosomes isolated from rye (Secale cereale L.) leaf chloroplasts were analyzed by two-dimensional polyacrylamide gel electrophoresis. The 50S subunit contained approx. 33 polypeptides in the range of relative molecular mass (M_r) 13000–36000, the 30S subunit contained approx. 25 polypeptides in the range of M_r 13000–40500. Antisera raised against the individual isolated ribosomal subunits detected approx. 17 polypeptides of the 50S and 10 polypeptides of the 30S subunit in the immunoblotting assay. By immunoblotting with these antisera the major antigenic ribosomal polypeptides (r-proteins) of the chloroplasts were clearly and specifically visualized also in separations of leaf extracts or soluble chloroplast supernatants. In extracts from rye leaves grown at 32° C, a temperature which is non-permissive for 70S-ribosome formation, or in supernatants from ribosome-deficient isolated plastids, six plastidic r-proteins were visualized by immunoblotting with the anti-50S-serum and two to four plastidic r-proteins were detected by immunoblotting with the anti-30S-serum, while other r-proteins that reacted with our antisera were missing. Those plastidic r-proteins that were present in 70S-ribosome-deficient leaves must represent individual unassembled ribosomal polypeptides that were synthesized on cytoplasmic 80S ribosomes. For the biogenesis of chloroplast ribosomes the mechanism of coordinate regulation appear to be less strict than those known for the biogenesis of bacterial ribosomes, thus allowing a marked accumulation of several unassembled ribosomal polypeptides of cytoplasmic origin.Abbreviations L polypeptide of large ribosomal subunit - M_r relative molecular mass - r-protein ribosomal polypeptide - S polypeptide of small ribosomal subunit - SDS sodium dodecyl sulfate     

UV-inducible proteins and UV-induced cross-protection against acid,ethanol, H2O2 or heat treatments in Lactococcus lactis subsp. lactis

The relationship between UV-irradiation-induced tolerance to different environmental stresses and change in protein synthesis was examined in Lactococcus lactis subsp lactis IL1403. The results showed that preirradiation of cultures of L. lactis subsp. lactis with UV_254nm light led to increased tolerance of usually lethal challenges to acid (pH 4.0), ethanol (20%, v/v), H₂O₂ (15 mM), or heat (52° C). This suggests that there is an overlapping regulation between the UV-induced pathway and the other stress responses. Whole-cell protein extracts from UV-treated (100 J/m²) and untreated cultures were compared using two-dimensional polyacrylamide gel electrophoresis. At least 14 polypeptides were induced in response to damage after UV irradiation, which indicated an SOS-like response in this species. The RecA protein, however, seemed not to be significantly induced in Lactococcus lactis subsp. lactis IL1403. Some of the UV-induced polypeptides overlaped with stress proteins induced by the other treatments.     

Iron Deficiency Induced Changes on Electron Transport Rate in Pisum Sativum Chloroplasts

Iron deficiency induced decrease in the rate of whole electron transport chain in chloroplasts of pea (Pisum sativum L.). Such reduction was mainly due to the loss of photosystem (PS) 2 activity. The same result was obtained when the ratio of variable to maximum chlorophyll fluorescence (F_v/F_m) was evaluated. The loss in PS 2 activity was primarily due to a loss of 33, 23 and 17 kDa polypeptides. In contrast, iron deficiency induced the synthesis of 28 and 29 kDa polypeptides.