Light-Harvesting Function in the Diatom Phaeodactylum tricornutum

Abstract

Three pigment-protein complexes were isolated from the marine dia-tom Phaeodactylum tricornutum (Bohlin) by treatment of thylakoid membrane fragments with 1% Triton X-100 at 4°C followed by centrifu-gation on sucrose density gradients. The major complex contains chlorophyll a, cl, C2, and the carotenoid fucoxanthin (chlorophyll a : c,: C2: fucoxanthin = 1.0: 0.09: 0.28 : 2.22) bound to an apoprotein doublet of 16.4 and 16.9 kilodaltons. This complex accounts for >70% of the total pigment and 20 to 40% of the protein in the thylakoid membranes. Efficient coupling of chlorophyll c and fucoxanthin absorption to chlorophyll a fluorescence supports a light-harvesting function for the complex. A minor light-harvesting complex containing chlorophyll a, cl, and C2 but no fucoxanthin (chlorophyll a: cl: C2 = 1.0: 0.23 : 0.26) was also isolated at Triton: chlorophyll a ratios between 20 and 40. These pigments are bound to a similar molecular weight apoprotein doublet. The third complex isolated was the P700-chlorophyll a protein, the reaction center of photosystem I, which showed characteristics similar to those isolated from other plant sources. The yield of the chlorophyll a/c-fucoxanthin complex was shown to respond strongly to changes in light intensity during growth, accounting for most of the changes in cellular pigmentation. The photosynthetically active pigments of plants are bound in discrete pigment-protein complexes (28) which can be divided functionally into two groups: the photochemical reaction center complexes where light energy is converted to chemical energy, and the light-harvesting complexes that serve as antennae to collect and transfer available light energy to the reaction centers. The P700-Chl a protein is the reaction center complex of PSI and is ubiquitous in all 02-evolving plants (28). Its pigment composition, structure, and spectral characteristics are probably highly conserved throughout the plant kingdom (29). The reaction center complex of PSII is not as well characterized as the P700-Chl a complex. Functional PSII complexes containing minimal amounts ofassociated antennae have been isolated (30). The majority of the Chl a and all of the accessory pigments (Chl b, c, photosynthetically active carotenoids and biliproteins) are bound in light-harvesting complexes which often aggregate into large macromolecular arrays in vivo. The LHCP2 of green 'Supported by grants to R. K. Clayton from the National Science Foundation (PCM-8202898) and the Department of Energy (DE-AC02-76ERO-3 162). 2Abbreviations. LHCP, light-harvesting Chl a/b protein; TB, 100 mm Tris borate (pH 8.0); bp, boiling point; LDS, lithium dodecyl sulfate; P700, reaction center pigment of PSI; Chl c, Chl c, + Chl c2. algae and higher plants and the phycobilisomes of red and blue-green algae are well characterized light-harvesting systems (9, 28). Both of these complexes deliver excitation energy preferentially to PSII and are probably involved in the regulation of excitation energy distribution between the two photosystems (12, 27). The diversity of pigments among the algal classes (16) and the importance of algae in global carbon fixation has stimulated the investigation of algal light-harvesting complexes. Chl cl and c2 are widely distributed in the nonchlorophyte eucaryotic algae and differ from Chl a and b in that they are unsaturated in Ring IV and are not esterified to a hydrocarbon alcohol (16). Chl and Chl-carotenoid complexes have been isolated from dinoflagel-lates (4, 24), cryptomonads (13), brown algae (1-3), and diatoms (7, 10). In most cases, the functional role of these complexes in light-harvesting was assessed by the efficiency of energy coupling between the accessory pigments and Chl a, but their respective roles in regulation of excitation energy distribution between the two photosystems has not been addressed. The aquatic light field differs in several respects from that in terrestrial habitats. Absorption and scattering processes by the medium as well as by dissolved and suspended materials rapidly attenuate total irradiance, and red and blue light in particular, with increasing depth (18). Far-red light (>680 nm) is particularly deficient at depths below the first few meters (18). This may present a problem to aquatic plants because light in the blue and far-red spectral regions are the only wavelengths oflight absorbed directly by PSI (19). Physical mixing processes create a temporal variability in the intensity and spectral distributions by transporting planktonic algae through the vertical light field on time scales of several h to several weeks. The response of cellular pigment content to variable light regimes has been described in most algal classes (23). In the present study, the isolation of the P700-Chl a protein and two functional light-harvesting complexes from the marine diatom Phaeodactylum tricornutum is reported. The isolation procedure utilizes solubilization of thylakoid membranes with 1% Triton X-100 followed by sucrose gradient centrifugation. The pigment and protein composition and spectral characteristics of the complexes were analyzed in terms of light absorption and the efficiency ofenergy transfer. The yield ofthe major light-harvesting complex is shown to change in response to vanrations in light intensity during growth. The role of the light-harvesting complexes in the regulation of excitation energy distribution between PSII and PSI is addressed in an accompanying report (20). MATERIALS AND METHODS Plant Material. Phaeodactylum tricornutum (Bohlin) was grown in unialgal axenic batch cultures at 12°C in f/2 enriched 732 https://plantphysiol.org Downloaded on May 20, 2021.-Published by