Fluorescence properties of chlorophyll a and b monomolecular films at the air-water interface

The fluorescence properties of chlorophyll $\text{a}$ and $\text{b}$ monomolecular films at the air-water interface were measured by a high sensitivity fluorophotometer using the photon-counting method. The fluorescence intensity of chlorophyll molecules in monomolecular films in the absence of any diluents did not decrease simply with the mean distance of chlorophyll molecules. Over the range of the mean distances from 27 to 21 Å, three fluorescence components (peaks at 685, 695 and 715 nm) of chlorophyll $\text{a}$ were observed. In the case of chlorophyll $\text{b}$, two fluorescence components (peaks at 667 and 685 nm) were observed over the range of the mean distances from 34 to 24 Å. When the mean distance was 18 Å, the short wavelength component of chlorophyll $\text{b}$ disappeared, and only the long wavelength component was observed.     

Antenna function of a chlorophyll ab protein complex of Photosystem I

The functional role of a chlorophyll $\text{a}\text{b}$ complex associated with Photosystem I (PS I) has been studied. The rate constant for P-700 photooxidation, K_P-700, which under light-limiting conditions is directly proportional to the size of the functional light-harvesting antenna, has been measured in two PS I preparations, one of which contains the chlorophyll $\text{a}\text{b}$ complex and the other lacking the complex. K_P-700 for the former preparation is half of that of the preparation which has the chlorophyll $\text{a}\text{b}$ complex present. This difference reflects a decrease in the functional light-harvesting antenna in the PS I complex devoid of the chlorophyll $\text{a}\text{b}$ complex. Experiments involving reconstitution of the chlorophyll $\text{a}\text{b}$ complex with the antenna-depleted PS I preparation indicate a substantial recovery of the K_P-700 rate. These results demonstrate that the chlorophyll $\text{a}\text{b}$ complex functions as a light-harvesting antenna in PS I.     

Specific activity of LHRH and TRH degrading enzymes in various tissues of normal and castrated male rats.

The chlorophyll composition and Hill activity of the leaf, developing seed parts and pod have been studied in three species of legumes, $\text{Lathyrus latifolius}$, $\text{Pisum sativum}$ and $\text{Vicia faba}$. The studies indicate that in all the species, the level of chlorophyll, on mg/g fresh weight basis, is maximum in the leaf. However, Hill activity studies show that the cotyledonary chloroplasts in all the cases have a higher Hill activity than the leaf chloroplasts. Thus, the Hill activities of the cotyledonary chloroplasts are 340% of the leaf chloroplasts in $\text{Lathyrus}$$\text{latifolius}$, 144% of the leaf chloroplasts in $\text{Pisum}$$\text{sativum}$ and 200% of the leaf chloroplasts in $\text{Vicia}$$\text{faba}$.     

Evidence for the localization of chlorophyll in lipid vesicles: a spin label study.

Fatty acid spin labels containing nitroxide groups at different positions in the fatty acid chain have been incorporated into lipid vesicles. Changes in esr parameters of the spin labels in the presence in the membrane of phytol, propionic acid phytol ester or chlorophyll $\text{a}$ and the kinetics of chlorophyll $\text{a}$ mediated photodestruction of the spin labels suggest a localization of the macrocyclic ring of the chlorophyll molecule in the polar head group region of the membrane.     

Isolation and characterization of Photosystem I and II membrane particles from the blue-green alga, Synechococcus cedrorum

Fractions enriched in either Photosystem I or Photosystem II activity have been isolated from the blue-green alga, Synechococcus cedrorum after digitonin treatment. Sedimentation of this homogenate on a 10–30% sucrose gradient yielded three green bands: the upper band was enriched in Photosystem II, the lowest band was enriched in Photosystem I, while the middle band contained both activities. Large quantities of both particles were isolated by zonal centrifugation, and the material was then further purified by chromatography on DEAE-cellulose.The resulting Photosystem II particles carried out light-induced electron transport from semicarbizide to ferricyanide of over 2000 μmol/mg Chlorophyll per h (which was sensitive to 3-(3,4-dichlorophenyl)-1,1-dimethylurea), and was nearly devoid of Photosystem I activity. This particle contains β-carotene, very little phycocyanin, has a chlorophyll absorption maximum at 675 nm, and a liquid N₂ fluorescence maximum at 685 nm. The purest Photosystem II particles have a chlorophyll to cytochrome $\text{b-559}$ ratio of 50 : 1. The Photosystem I particle is highly enriched in $\text{P-700}$, with a chlorophyll to $\text{P-700}$ ratio of 40 : 1. The physical structure of the two Photosystem particles has also been studied by gel electrophoresis and electron microscopy. These results indicate that the size and protein composition of the two particles are distinctly different.     

Isolation of the light-harvesting chlorophyll a/b--protein complex from thylakoid membranes of barley by adsorption chromatography on controlled-pore glass.

The light-harvesting chlorophyll $\text{a}\text{b}$-protein complex has been isolated from barley thylakoids by a rapid, single-step procedure involving adsorption chromatography on controlled-pore glass columns. The Triton X-100-solubilized complex contains a polypeptide of apparent molecular weight, 26,000; the 0.25% Triton X-100 light-harvesting chlorophyll $\text{a}\text{b}$-protein has spectral characteristics consistent with its assumed in vivo state. On the same column free chlorophyll and carotenoids have been separated from chlorophyll-protein complex 1, but this complex contained many polypeptides other than those associated with chlorophyll. This method is potentially suitable for the isolation of other thylakoid membrane proteins. It may also be generally applicable for fractionation of intrinsic membrane proteins from other sources and for separation of mixed Triton X-100-lipid micelles.     

Light-harvesting systems of brown algae and diatoms. Isolation and characterization of chlorophyll ac and chlorophyll afucoxanthin pigment-protein complexes

The present study examined the protein associations and energy transfer characteristics of chlorophyll c and fucoxanthin which are the major light-harvesting pigments in the brown and diatomaceous algae. It was demonstrated that sodium dodecyl sulfate (SDS)-solubilized photosynthetic membranes of these species when subjected to SDS polyacrylamide gel electrophoresis yielded three spectrally distinct pigment-protein complexes. The slowest migrating zone was identical to complex I, the SDS-altered form of the P-700 chlorophyll a-protein. The zone of intermediate mobility contained chlorophyll c and chlorophyll a in a molar ratio of 2 : 1, possessed no fucoxanthin, and showed efficient energy transfer from chlorophyll c to chlorophyll a. The fastest migrating pigment-protein zone contained fucoxanthin and chlorophyll a, possessed no chlorophyll c, and showed efficient energy transfer from fucoxanthin to chlorophyll a. It is demonstrated that the chlorophyll $\text{a}\text{c-}\text{protein}$ and the chlorophyll $\text{a}\text{fucoxanthin-protein}$ complexes are common to the brown algae and diatoms examined, and likely share similar roles in the photosynthetic units of these species.     

Low-intensity subnanosecond fluorescence study of the light-harvesting chlorophyll ab protein

The fluorescence decay characteristics of the isolated light-harvesting chlorophyll $\text{a}\text{b}$ protein have been studied using low-intensity subnanosecond-resolution time-correlated single-photon counting. In the monomeric state in detergent micelles, the chlorophyll $\text{a}\text{b}$ protein exhibits biexponential decay (τ₁ = 1.2 ns, τ₂ = 3.3 ns) with the two components having very similar weights. The decay parameters do not depend on emission wavelength. These results are discussed in relation to the Van Metter-Knox-Shepanski model (Van Metter, R.M. (1977) Biochim. Biophys. Acta 462, 642–657; Shepanski, J.S. and Knox, R.S. (1982) Isr. J. Chem., in the press) of the chlorophyll $\text{a}\text{b}$ protein, and a kinetic analysis of the energy-transfer processes. The influence of detergent composition and concentration on the fluorescence decay of the chlorophyll protein is also described.     

Zero field resonance and spin alignment of the triplet state of chloroplasts at 2°K

Microwave induced transitions in zero magnetic field have been observed in the photoinduced triplet of chloroplasts treated with dithionite by monitoring changes in the intensity of the 735 nm fluorescence band at 2°K. Similar results were obtained with chloroplasts treated with hydroxylamine plus 3-(3,4-dichlorophenyl)-1,1-dimethylurea and preillumination. The zero field parameters are $\text{D = 0.02794 ± 0.00007}\text{cm}{}^{\mathrm{?1}}$, $\text{E = 0.00382 ± 0.00007}\text{cm}{}^{\mathrm{?1}}$, i.e. equal to those of monomeric chlorophyll $\text{a}$ to within the experimental error. The photoinduced triplet appears to be linked to Photosystem II. This indicates that the low temperature 735 nm fluorescence band of chloroplasts is at least partly due to Photosystem II.     

Isolation and characterization of a major chlorophyll ac2 light-harvesting protein from a Chroomonas species (Cryptophyceae)

Three chlorophyll-protein complexes of a Chroomonas species (Cryptophyceae) have been separated by sodium dodecyl sulphate polyacrylamide gel electrophoresis. The two bands at 100 and 42 kDa are Complex I (CP I) and Complex IV (CP IV), the ubiquitous chlorophyll a-proteins associated with Photosystems I and II, respectively. The third 55 kDa band, which had two peptide subunits (24 and 20 kDa), contained both chlorophyll a and chlorophyll c₂ in a molar ratio of 1.4 chlorophyll a to 1 chlorophyll c₂ ($\text{chlorophyll}\text{a}\text{chlorophyll}\text{c}{}_2$ ratio in whole cells = 4). A chlorophyll $\text{a}\text{c}{}_2$ fraction with similar spectral and electrophoretic properties was isolated by digitonin-sucrose density gradient centrifugation. This fraction had no photochemical activity and contained only a single carotenoid species with absorbance maxima in methanol at 424, 448 and 476 nm. Efficient energy transfer from chlorophyll c₂ to chlorophyll a occurred in the complex.     

Changes in composition and function of thylakoid membranes as a result of photosynthetic adaptation of chloroplasts from pea plants grown under different light conditions

The hypothesis that chloroplasts having different light-saturated rates of photosynthesis will have different proportions of the intrinsic thylakoid complexes engaged in light-harvesting and electron transport (Anderson, J.M. (1982) Mol. Cell. Biochem. 46, 161–172) has been tested. Peas were grown in light regimes which varied in light intensity, quality and time of irradiance, and ranged from sunlight through red to blue-enriched light of very low radiation. The electron-transport capacity at saturating light of Photosystem I and Photosystem II of chloroplasts isolated from light-adapted peas was 2-fold and 5–6-fold lower, respectively, in the lowest radiation compared to sunlight. There was a marked increase in the amount of total chlorophyll associated with the main chlorophyll $\text{a}\text{b-}\text{proteins}$ (LHCP¹, LHCP² and LHCP³) and a 2-fold decrease in the core reaction centre complex of Photosystem II (CP a) as the radiation decreased; the $\text{LHCP}{}^{\mathrm{1–3}}\text{CP}$ a ratio changed from 3.5 to 9.0. The amount of chlorophyll associated with Photosystem I varied from 34% in sunlight to 27% in the lowest radiation, but the antenna size of Photosystem I was not markedly different; there was a 2-fold decrease in the amount of cytochrome f on a chlorophyll basis, which partly accounted for the decreased electron-transport capacity of Photosystem I. Since the increases or decreases in the levels of each of the components correlated with decreasing radiation, it is clear that the light-adaptation of both light-harvesting and electron-transport components is indeed closely co-ordinated.     

Picosecond time-resolved study of MgCl2-induced chlorophyll fluorescence yield changes from chloroplasts

The MgCl₂-induced chlorophyll fluorescence yield changes in broken chloroplasts, suspended in a cation-free medium, treated with 3,-(3′,4′-dichlorophenyl)-1,1-dimethylurea and pre-illuminated, has been investigated on a picosecond time scale. Chloroplasts in the low fluorescing state showed a fluorescence decay law of the form exp $\text{?At}{}^{\mathrm{<mtext>1</mtext><mtext>2</mtext>}}$, where A was found to be 0.052 $\text{ps}{}^{\mathrm{<mtext>?1</mtext><mtext>2</mtext>}}$, and may be attributed to the rate of spillover from Photosystem II to Photosystem I. Addition of 10 mM MgCl₂ produced a 50% increase in the steady-state fluorescence quantum yield and caused a marked decrease in the decay rate. The fluorescence decay law was found to be predominantly exponential with a 1/e lifetime of 1.6 ns. These results support the hypothesis that cation-induced changes in the fluorescence yield of chlorophyll are related to the variations in the rate of energy transfer from Photosystem II to Photosystem I, rather than to changes in the partitioning of absorbed quanta between the two systems.     

Temperature dependence of the magnetic susceptibility of cobalt (II) stellacyanin.

Magnetic susceptibility measurements on cobalt(II) stellacyanin ($\text{Rhus vernicifera}$) have been performed between 2.2 and 50 K. The effective magnetic moment of Co(II) in the protein is 3.91 ± 0.12 (μ_B). Nonlinear behavior below 3 K evidences the presence of zero-field splitting attributable to a low-symmetry component of the ligand field. The results are consistent with a structural model based on a distorted tetrahedral Co(II) site involving one or more extremely covalent metal-ligand bonds.     

Chlorophyll-protein complexes of a Codium species,including a light-harvesting siphonaxanthin-Chlorophylla ab-protein complex,an evolutionary relic of some Chlorophyta

Eight chlorophyll-protein complexes were isolated from thylakoid membranes of a Codium species, a marine green alga, by mild SDS-polyacrylamide gel electrophoresis. CP 1a¹, CP 1a², CP 1a³ and CP 1a⁴ were partially dissociated Photosystem (PS) I complexes, which in addition to the core reaction centre complex, CP 1, possessed PS I light-harvesting complexes containing chlorophyll (Chl) a, Chl b and siphonaxanthin. LHCP¹ and LHCP³ are orange-brown green chlorophyll $\text{a}\text{b-}\text{proteins}$ ($\text{Chl}\text{a}\text{Chl}\text{b}$ ratios of 0.66) that contain siphonaxanthin and its esterified form, siphonein. CP a and CP 1, the core reaction centre complexes of PS II and PS I, respectively, had similar spectral properties to those isolated from other algae or higher plants. These P-680- or P-700-Chl a-proteins are universally distributed among algae and terrestrial plants; they appear to be highly conserved and have undergone little evolutionary adaptation. Siphonaxanthin and siphonein which are present in the Codium light-harvesting complexes of PS II and PS I are responsible for enhanced absorption in the green region (518 and 538 nm). Efficient energy transfer from both xanthophylls and Chl b to only Chl a in Codium light-harvesting complexes, which have identical fluorescence emission spectra at 77 K to those of the lutein-Chl $\text{a}\text{b-}\text{proteins}$ ($\text{Chl}\text{a}\text{Chl}\text{b}$ ratios of 1.2) of most green algae and all higher plants, proved that the molecular arrangement of these light-harvesting pigments was maintained in the isolated Codium complexes. The siphonaxanthin-Chl $\text{a}\text{b-}\text{proteins}$ allow enhanced absorption of blue-green and green light, the predominant light available in deep ocean waters or shaded subtidal marine habitats. Since there is a variable distribution of lutein, siphonaxanthin and siphonein in marine green algae and siphonaxanthin is found in very ancient algae, these novel siphonein-siphonaxanthin-Chl $\text{a}\text{b-}\text{proteins}$ may be ancient light-harvesting complexes which were evolved in deep water algae.     

Inhibition of cyclopropane fatty acid synthase by sinefungin and A9145C

Sinefungin and A9145C, antifungal antibiotic analogs of S-adeno-sylmethionine isolated from $\text{Streptomyces}\text{,}\text{griseolus}$, have been found to be very effective $\text{in}\text{,}\text{vitro}$ inhibitors of cyclopropane fatty acid synthase from $\text{Lactobacillus}\text{,}\text{plantarum}$. Both compounds exhibit linear competitive inhibition with a K_i for Sinefungin of 220 nM and a K_i for A9145C of 11 nM.     

Resolution of the light-harvesting chlorophyll ab-protein of Vicia faba chloroplasts into two different chlorophyll-protein complexes

Thylakoids of Vicia faba chloroplasts disaggregated by sodium dodecyl sulfate were separated by means of different electrophoretic systems. Under the conditions of a high resolving gel system the chlorophyll containing zone previously termed chlorophyll-protein complex II or light-harvesting chlorophyll $\text{a}\text{b}\text{-}\text{protein}$ was found to be inhomogeneous. It represents a mixture of two distinct chlorophyll-proteins characterized by different spectral properties and different apoproteins. One chlorophyll-protein exhibits a chlorophyll $\text{a}\text{b}$ ratio of 0.9 and is associated with polypeptides of 24 000 and 23 000 daltons. The 24 000 dalton band is proved to bind chlorophyll and has a light-harvesting function. The function of the 23 000 dalton band is unknown. The second chlorophyll-protein has a chlorophyll $\text{a}\text{b}$ ratio of 2.1 and an additional absorption maximum in the position of 637 nm. It is associated with only one polypeptide which has an apparent molecular weight of 23 000. The two 23 000 dalton polypeptides occurring in both complexes are not identical.     

Hydroxylation of beta-phenylethylamine in the rat

$\text{ortho}$, $\text{meta}$ and $\text{para}$ Tyramine, as their dansyl derivatives, have been identified and quantitated by mass spectrometry in rat urine. After intraperitoneal injection of phenylethylamine labelled either with deuterium or tritium in the presence of pargyline, the amount of urinary tyramines excreted was 0.8% ($\text{para}$, 0.74%; $\text{meta}$, 0.12%; $\text{ortho}$, 0.008%) of the injected dose. The advantages of involving stable isotopes and mass spectrometry in metabolic studies of pharmacologically active compounds is discussed.     

The nature of the light-harvesting complex as defined by sodium dodecyl sulfate polyacrylamide gel electrophoresis

Reelectrophoresis of the oligomer form (CP II1) of the chlorophyll $\text{a}\text{b}$ light-harvesting complex (LHC) from the green alga Acetabularia yields two green bands which run at the position typical of the monomer (CP II). The upper green band (CP II₁) is enriched in the 27 kDa polypeptide of the LHC, while the lower is enriched in the 26 kDa polypeptide. The fact that both bands have both chlorophyll (Chl) a and b, and in the same ratio, implies that the LHC is made up of two Chl $\text{a}\text{b}$ proteins. Neither of these bands can be attributed to the Chl $\text{a}\text{b}$ complex ‘CP 29’ (Camm, E.L. and Green, B.R. (1980) Plant Physiol. 66, 428–432). Resolution of CP II₁ and CP II₂ of spinach can be obtained if sucrose gradient fractions of an octylglucoside extract are subjected to SDS-polyacrylamide gel electrophoresis. CP II₁ and CP II₂ are interpreted as being fundamental subunits of the light-harvesting complex as it is defined on SDS-polyacrylamide gels.     

Letter: Electron microscopy of pyruvate kinase crystals from Saccharomyces carlsbergensis

Crystals of pyruvate kinase have been analysed by electron microscopy, optical diffraction and filtering, and the following parameters were obtained: $\text{2a = 93}\text{A}\text{?}$, $\text{2b = 126}\text{A}\text{?}$, $\text{l = 35·2}\text{A}\text{?}$. A comparison of these data with the parameters obtained from small-angle X-ray scattering measurements indicates that two molecules of the tetrameric enzyme are arranged in one packing unit.