Chlorophyll‐protein complexes of barley photosystem I

Abstract

Photosystem I (PSI) preparations with a chlorophyll a/b ratio of 6.0 were isolated from barley thylakoids using two different methods. The high-molecular-mass complex (CPla*) which is resolved by non-denaturing gel electrophoresis had the same properties as a PSI preparation (PSI-200) isolated by Triton X-1 00 solubilisation of thylakoids followed by sucrose gradient ultracentrifugation. This material had a chlorophyll : P700 ratio of 208 : I and was composed of three different chlorophyll-protein complexes which could be separated from each other by solubilising the PSI preparation in dodecyl maltoside followed by sucrose gradient ultracentrifugation. Approximately half of the chlorophyll, including all the chlorophyll h, was located in two antenna complexes designated LHCI-680 and LHCI-730, which were identified by their characteristic low-temperature fluorescence emission spectra. The rest of the chlorophyll a was associated with the PSI reaction centre, P700 Ch1,-P1, which fluoresced at 720 nm. Each chlorophyll-protein complex had a unique polypeptide composition and characteristic circular dichroic and absorption spectra. The use of dodecyl maltoside instead of dodecyl sulphate resulted in a less denatured form of LHCI-680, which fluoresced at 690 nm at 77 K. One of the sucrose gradient fractions contained a complex consisting of only LHCI-730 and P700 Ch1,-P1 which fluoresced at 731 nm, indicating that LHCI-730 is structurally associated with P700 Chl,-P3 and quenches its fluorescence. Approximately threequarters of the light-harvesting antenna chlorophyll was in LHCI-730, but only about one-quarter of the normal complement of LHCI-730 was required to quench the reaction centre.By reducing the amount of Triton relative to the chlorophyll concentration, a PSI preparation (chlorophyll a/h ratio of 3.5) with a chlorophyll:P700 ratio of 300:l was isolated. It contained no photosystem 11, but a significant amount of LHCII which was functionally connected to the PSI reaction centre. Reconstitution studies demonstrated that excitation energy transfer from LHCII to PSI requires the presence of LHCI-680, and we propose that, in PSI, the following linear excitation energy transfer sequence occurs: LHCII + LHCI-680 4 LHCI-730 + P700 Chl,-Pl.Much of our present knowledge of the structure and composition of photosystem I (PSI) in higher plant thylakoids was made possible by the work of Mullet et al. [l] who isolated and characterised a functional PSI preparation in high yield from pea leaves. It retained the long-wavelength low-temperature fluorescence emission typical of whole thylakoids and supported electron transport from dichloroindophenol to methyl viologen. The existence of a PSI-specific lightharvesting chlorophyll-protein complex (LHCI) was predicted from the reduction in antenna size of a PSI 'core' preparation [l] Abbreviations. CD, circular dichroism; Chl, chlorophyll; LHC, light-harvesting chlorophyll protein; PAGE, polyacrylamide gel electrophoresis; PS, photosystem; SDS, sodium dodecyl sulphate. fluores...

Protein phosphorylation and Mg2+influence light harvesting and electron transport in chloroplast thylakoid membrane material containing only the chlorophyll‐a/b‐binding light‐harvesting complex of photosystem II and photosystem I

“…The reaction-centre polypeptide of PSI is clearly present in the PSI/LHC-I1 material as a monomer with an approximate molecular mass of 65 kDa and as a dimer of approximate molecular mass 130 kDa. LHC-I polypeptides in the mass range 25 -20 kDa are also evident [24,331, indicating that the material represents an LHC-11/ LHC-I/PSI reaction-centre association. It is clear that LHC-I1 polypeptides in the range 28-25 kDa make up a large proportion of the overall assembly.…”

“…This material, similar in polypeptide composition to a complex isolated from barley [24], has been shown by freezefracture electron microscopy to be composed of crystalline sheets of particles approximately 8 nm in diameter and interspersed with larger particles of diameter 13-17 nm [23]. The ordered sheets of 8-nm particles were similar in morphology to the 2-dimensional crystals of purified LHC-I1 [25].…”

“…If PSI centres are associated with 210 molecules of chlorophyll u and b, in a ratio of 6.0 [24] and LHC-I1 binds 13-15 chlorophylls u and hlpolypeptide in a ratio of 1.1 [34], then a LHC-II/PSI polypeptide stoichiometry of 12 can be calculated for an assembly containing PSI and LHC-11 and having a chlorophyll u/b ratio of 2.6. This would equate to a LHC-11/ PSI stoichiometry of 4 when the trimeric assembly of LHC-I1 is considered.…”

Probing in vitro translation products with monoclonal antibodies to chlorophylla/b-binding proteins of barley thylakoids

“…What the change in mobility is due to is not clear, but it is always observed (4,14). Figure 2 shows that CMpLHCI:2 reacts with the Thylakoid polypeptides from wild-type seedlings (20 lag Chl) illuminated for 6, 9 or 12 hours were separated on 11-15% SDS-PAGE prior to electroblotting.…”

“…In barley photosystem I (PSI) preparations isolated by Triton X-100 solubilisation of thylakoids yield upon sucrose gradient centrifugation two chlorophyll a/b-binding proteins. They are identified by their low-temperature fluorescence emission spectra and are named LHCI-680 and LHCI-730 (2,4,17). Photosystem II (PSII)…”