Michael D. Spangfort

A nomenclature for the genes encoding the chlorophylla/b-binding proteins of higher plants

We propose a nomenclature for the genes encoding the chlorophylla/b-binding proteins of the light-harvesting complexes of photosystem I and II. The genes encoding LHC I and LHC II polypeptides are namedLhca1 throughLhca4 andLhcb1 throughLhcb6, respectively. The proposal follows the general format recommended by the Commision on Plant Gene Nomenclature. We also present a table for the conversion of old gene names to the new nomenclature.

Chlorophyll a/b-Binding Proteins, Pigment Conversions, and Early Light-Induced Proteins in a Chlorophyll b-less Barley Mutant

Monospecific polyclonal antibodies have been raised against synthetic peptides derived from the primary sequences from different plant light-harvesting Chl a/b-binding (LHC) proteins. Together with other monospecific antibodies, these were used to quantify the levels of the 10 different LHC proteins in wild-type and chlorina f2 barley (Hordeum vulgare L.), grown under normal and intermittent light (ImL). Chlorina f2, grown under normal light, lacked Lhcb1 (type I LHC II) and Lhcb6 (CP24) and had reduced amounts of Lhcb2, Lhcb3 (types II and III LHC II), and Lhcb4 (CP 29). Chlorina f2 grown under ImL lacked all LHC proteins, whereas wild-type ImL plants contained Lhcb5 (CP 26) and a small amount of Lhcb2. The Chlorina f2 ImL thylakoids were organized in large parallel arrays, but wild-type ImL thylakoids had appressed regions, indicating a possible role for Lhcb5 in grana stacking. Chlorina f2 grown under ImL contained considerable amounts of violaxanthin (2-3/reaction center), representing a pool of phototransformable xanthophyll cycle pigments not associated with LHC proteins. Chlorina f2 and the plants grown under ImL also contained early light-induced proteins (ELIPs) as monitored by western blotting. The levels of both ELIPs and xanthophyll cycle pigments increased during a 1 h of high light treatment, without accumulation of LHC proteins. These data are consistent with the hypothesis that ELIPs are pigment-binding proteins, and we suggest that ELIPs bind photoconvertible xanthophylls and replace “normal” LHC proteins under conditions of light stress.

LIGHT-ABSORPTION AND ELECTRON-TRANSPORT BALANCE BETWEEN PHOTOSYSTEM II AND PHOTOSYSTEM I IN SPINACH CHLOROPLASTS

Abstract— The distribution of absorbed light and the turnover of electrons by the two photosystems in spinach chloroplasts was investigated. This was implemented upon quantitation of photochemical reaction centers, chlorophyll antenna size and composition of each photosystem (PS), and rate of light absorption in situ. In spinach chloroplasts, the photosystem stoichiometry was PSIIJPSIIα/PSIIβ/PSI= 1.3/0.4/1.0. The number (N) of chlorophyll (a+b) molecules associated with each PS was N(PSIIα)/N(PSIIβ)/N(PSI)=230/100/200, i.e. about 65% of all Chl is associated with PSII and about 35% with PSI. Light absorption by PSII in vivo is selectively attenuated at the molecular, membrane and leaf levels, (a) The rate of light absorption by PSII was only 0.85 that of PSI because of the lower rate of light absorption by Chl b as compared to Chl a (approximately 80% of all Chl b in the chloroplast is associated with PSII). (b) The exclusive localization of PSIIα in the membrane of the grana partition regions and of PSI in intergrana lamellae resulted in a differential “sieve effect” or “flattening of absorbance” by the photosystems in the two membrane regions. Due to this phenomenon, the rate of light absorption by PSII was lower than that of PSI by 15‐20%. (c) Selective filtering of sunlight through the spinach leaf results in a substantial distortion of the effective absorbance spectra and concomitant attenuation of light absorption by the two photosystems. Such attenuation was greater for PSII than for PSI because the latter benefits from light absorption in the 700‐730 nm region.

Phosphorylation controls the three-dimensional structure of plant light harvesting complex II.

The most abundant chlorophyll-binding complex in plants is the intrinsic membrane protein light-harvesting complex II (LHC II). LHC II acts as a light-harvesting antenna and has an important role in the distribution of absorbed energy between the two photosystems of photosynthesis. We used spectroscopic techniques to study a synthetic peptide with identical sequence to the LHC IIb N terminus found in pea, with and without the phosphorylated Thr at the 5th amino acid residue, and to study both forms of the native full-length protein. Our results show that the N terminus of LHC II changes structure upon phosphorylation and that the structural change resembles that of rabbit glycogen phosphorylase, one of the few phosphoproteins where both phosphorylated and non-phosphorylated structures have been solved. Our results indicate that phosphorylation of membrane proteins may regulate their function through structural protein-protein interactions in surface-exposed domains.

Isolation and characterization of the chlorophyll a/b protein complex CP29 from spinach

We describe a preparative isolation method for a minor chlorophyll a/b protein complex from spinach Photosystem II, which we suggest is identical to the CP29 complex first reported by Machold, O. and Meister, A. (Biochim. Biophys. Acta 546 (1979) 472–480) and by Camm, E.L. and Green, B.R. (Plant Physiol. 66 (1980) 428–432). The CP29 complex was isolated in mg quantities by ion-exchange high-performance liquid chromatography and extensively characterized. The absorption maximum in the red region was at 678 nm. The purified complex retained 10–12 chlorophyll molecules per protein and had a chlorophyll a/b ratio of 3.0–3.2. The complex contained only one apoprotein with an apparent molecular weight between 29 and 31.5 kDa. It showed a fluorescence emission peak at 679 nm when measured at 77 K while the fluorescence excitation spectrum revealed three main peaks in the blue region at 431 nm (chlorophyll a ), 468 nm (chlorophyll b ) and 493 nm (carotenoids). N-terminal sequencing of fragments from the trypsin-treated CP29 gave a consensus sequence of 29 amino acids.

Energy transfer within the isolated light‐harvesting chlorophyll a/b protein of photosystem II (LHC‐II)

Picosecond absorption spectroscopy has been used to study energy transfer within the isolated light‐harvesting chlorophyll a/b protein complex of photosystem II (LHC‐II). The results suggest: (i) a fast transfer (τ = 6 ± 4 ps) from chlorophyll b to chlorophyll a, (ii) during the lifetime of the chlorophyll b excited state there is no transfer of excitation energy between differently oriented molecules of chlorophyll b, (iii) a fast redistribution (τ = 20 ps) of excitation energy between different chlorophyll a chromophores.

Ultrafast chlorophyll b-chlorophyll a excitation energy transfer in the isolated light harvesting complex, LHC II, of green plants: Implications for the organisation of chlorophylls

The excitation energy transfer between chlorophyll b (Chl b) and chlorophyll a (Chl a) in the isolated trimeric chlorophyll‐a/b‐binding protein complex of spinach photosystem 2 (LHC II) has been studied by femtosecond spectroscopy. In the main absorption band of Chl b the ground state recovery consists of two components of 0.5 ps and 2.0 ps, respectively. Also in the Chl a absorption band, at 665 nm, the ground state recovery is essentially bi‐exponential. In this case is, however, the fastest relaxation lifetime is a 2.0 ps component followed by a slower component with a lifetime in the order of 10–20 ps. In the Chl b absorption band a more or less constant anisotropy of r = 0.2 was observed during the 3 ps the system was monitored. In the Chl a absorption band there was, however, a relaxation of the anisiotropy from r = 0.3 to a quasi steady state level of r = 0.18 in about 1 ps. Since the 0.5 ps component is only seen upon selective excitation of Chl b we assign this component to the energy transfer between Chl b and Chl a. The other components most likely represents redistribution processes of energy among spectrally different forms of Chl a. The energy transfer process between Chl b and Chl a can well be explained by the Förster mechanism which also gives a calculated distance of 13 Å between interacting chromophores. The organisation of chlorophylls in LHC II is discussed in view of the recent crystal structure data (1991) Nature 350, 130].

Epitope grafting: re-creating a conformational bet V 1 antibody epitope on the surface of the homologous apple allergen MAL D 1

Birch-allergic patients often experience oral allergy syndrome upon ingestion of vegetables and fruits, most prominently apple, that is caused by antibody cross-reactivity of the IgE antibodies in patients to proteins sharing molecular surface structures with the major birch pollen group 1 allergen from Betula verrucosa (Bet v 1). Still, to what extent two molecular surfaces need to be similar for clinically relevant antibody cross-reactivity to occur is unknown. Here, we describe the grafting of a defined conformational antibody epitope from Bet v 1 onto the surface of the homologous apple allergen Malus domestica group 1 (Mal d 1). Engineering of the epitope was accomplished by genetic engineering substituting amino acid residues in Mal d 1 differing between Bet v 1 and Mal d 1 within the epitope defined by the mAb BV16. The kinetic parameters characterizing the antibody binding interaction to Bet v 1 and to the mutated Mal d 1 variant, respectively, were assessed by Biacore experiments demonstrating indistinguishable binding kinetics. This demonstrates that a conformational epitope defined by a high affinity antibody-allergen interaction can successfully be grafted onto a homologous scaffold molecule without loss of epitope functionality. Furthermore, we show that increasing surface similarity to Bet v 1 of Mal d 1 variants by substitution of 6–8 residues increased the ability to trigger basophil histamine release with blood from birch-allergic patients not responding to natural Mal d 1. Conversely, reducing surface similarity to Bet v 1 of a Mal d 1 variant by substitution of three residues abolished histamine release in one patient reacting to Mal d 1.

Excitation energy annihilation in aggregates of chlorophyll ab complexes

Time-resolved picosecond absorption measurements were performed on aggregates of the light-harvesting chlorophyll (Chl) ab complexes from spinach thylakoids. Variation of the intensity of the exciting laser pulse showed that efficient excitation annihilation occurs in these aggregates. From the time-integrated absorption decays, the domain size of these aggregates was calculated to be at least 300 and probably about 1000 chlorophyll a molecules. From the time-resolved chlorophyll a excited state decays, the rate of excitation annihilation per pair of excitations, γ2, was calculated to be (2–3)·109s−1. It was shown that the annihilation competes with a 400 ps decay component in the mono-excitation decay, probably due to Chl ab aggregates. A second decay phase of 2–3 ns is not associated with annihilation. The calculated domain size and annihilation rate constants correspond to a nearest-neighbour transfer rate of at least 2·1011s−1 but probably closer to 1012s−1. Our results show that although these Chl ab complexes are dissociated from the membrane, their aggregation is such that fast energy transfer occurs over a large number of chlorophyll molecules.

Indoor allergen levels in Guangzhou city, southern China

To cite this article: Zhang C, Gjesing B, Lai X, Li J, Spangfort MD, Zhong N. Indoor allergen levels in Guangzhou city, southern China. Allergy 2011; 66: 186–191.