Interaction between phycobilisomes from Porphyridium cruentum and thylakoid membranes from Gymnodinium sp or spinach

Thylakoid membranes were isolated from Gymnodinium sp. and spinach, whereas the phycobilisomes were isolated and purified from red alga Porphyridium cruentum. The absorption spectra of the purified phycobilisomes (PBS) showed three peaks at 548, 564, and 624 nm, respectively, and the ratio of the fluorescence intensity at the lambda(680)(em) to lambda(80)(em5) that at was about 7.3. All these results demonstrated that the purified PBS remained intact. The thylakoid membranes were incubated with the purified phycobilisomes, and the thylakoid membranes, which harbored the phycobilisomes, were purified by sucrose density gradient centrifugation. Meantime, the conjugates of phycobilisome-thylakoid membranes were constructed using glutaraldehyde and further purified. Their characteristics were studied by measuring the absorption spectra and fluorescence emission spectra. The results showed that the phycobilisomes from Porphyridium cruentum can attach to the thylakoid membranes from Gymnodinium sp. and spinach without covalent cross-linking, but the excited energy transfer did not occur. The conjugate of phycobilisome-thylakoid. membranes with covalent cross-linking exhibits the excited energy transfer between the phycobilisomes and the thylakoid membranes.

Characterization of the main light-harvesting chlorophyll a/b-protein complex of green alga, Bryopsis corticulans

The main light-harvesting chlorophyll a/b -protein complex (LHC II) has been isolated directly from thylakoid membranes of shiphonous green alga, Bryopsis corticulans Setch. by using two consecutive runs of anion exchange and gel-filtration chromatography. Monomeric and trimeric subcomplexes of LHC 11 were obtained by using sucrose gradient ultracentrifugation. Pigment analysis by reversed-phase high performance liquid chromatography showed that chlorophyll a (Chl a), chlorophyll b (Chl b), neoxanthin, violaxanthin and siphonaxanthin were involved in LHC 11 from B. corticulans. The properties of electronic transition of monomeric LHC II showed similarities to those of trimeric LHC II. Circular dichroism spectroscopy showed that strong intramolecular interaction of excitonic dipoles between Chl a and between Chl b exist in one LHC II apoprotein, while the intermolecular interaction of these dipoles can be intensified in the trimeric structure. The monomer has high efficient energy transfer from Chl b and siphonaxanthin to Chl a similarly to that of the trimer. Our results suggest that in B. corticulans, LHC II monomer has high ordered pigment organization that play effective physiological function as the trimer, and thus it might be also a functional organization existing in thylakoid membrane of B. corticulans.

The stabilization effect of glutaraldehyde on the Spirulina plateusis phycobilisomes

The spectral properties of the glutaraldehyde-treated phycobilisomes were studied. The results showed that glutaraldehyde was effective in preventing phycobilisomes from dilution-induced dissociation and preserving the intra-phycobilisomes energy transfer.

Characterization of the artificially covalent conjugate of B-phycoerythrin and R-phycocyanin and the phycobilisome from Porphyridium cruentum

B-phycoerythrin (BPE) and R-phycocyanin (RPC) were purified from Porphyridium cruentum by Sephadex G-200 chromatography, then the BPE was attached covalently to the RPC by reacting their amino groups to form the artificially covalent BPE-RPC conjugate in which the excitation energy can transfer from the BPE to the RPC with low efficiency. Meanwhile, the intact phycobilisome (PBS) consisting of BPE, RPC, APC and L-CM was isolated and purified from Porphyridium cruentum, and the purified PBS was found to keep intact if the solution contains sucrose. Comparison of spectroscopic properties between the purified PBS and the BPE-RPC conjugate suggests that the BPE-RPC conjugate is much more stable than the purified PBS. The construction of BPE-RPC conjugate with low efficiency of the excitation energy transfer may be useful for preparing phycobiliprotein probes. (C) 2002 Elsevier Science Ireland Ltd. All rights reserved.

Isolation of PSI from a siphonous marine green alga, Codium fragile

Three different forms of PS I complexes were isolated from a siphonous marine green alga, Codium fragile, by Triton X-100 sucrose gradient centrifugation. Zone III had a Chl a/b>20, and designated as PS I. core complex CC I because it created only CP I band in mild PAGE. Zone IV and V had absorption at 436 and 674 nm, 467 and 650 nm, and 540 nm, suggesting the presence of Chl a, Chl b, siphonaxanthin and siphonein, Chl a/b were 3.23 and 2.4, respectively. Both CP I and CP I a bands were observed when they were subjected to mild PAGE. Therefore, Zone IV and V were different forms of PS I complexes that consisted of CC I and different amount of light-harvesting complex LHC I. Zone III contained only 66 and 56 ku peptides in SDS-PAGE, while Zone IV and V had 4 different LHC I peptides of 25, 26, 26.2 and 27.5 ku in addition to 66, 56 ku peptides. Fluorescence emission spectra showed that efficient energy transfer were kept among pigments in isolated PS I complexes. Excitation energy absorbed by Chl b, siphonaxanthin and siphonein can be transferred to Chl a.

Characteristics and comparative study of chlorophyll-protein complexes from siphonous green algae

By mild PAGE method, 11, 11, 7 and 9 chlorophyll-protein complexes were isolated from two species of siphonous green algae ( Codium fragile (Sur.) Harlot and Bryopsis corticulans Setch.), green alga (Ulothrix flacca (Dillw.) Thur.), and spinach (Spinacia oleracea Mill.), respectively. Apparent molecular weights, Chi a/b ratios, distribution of chlorophyll, absorption spectra, low temperature fluorescence spectra of these complexes were determined, and compared with one another. PS I complexes of two siphonous green algae are larger in apparent molecular weight because of the attachment of relative highly aggregated LHC I. Four isolated light-harvesting complexes of PSII are all siphonaxanthin-Chl a/b-protein complexes, and they are not monomers and oligomers like those in higher plants. Especially, the absence of 730 nn fluorescence in PS I complexes indicates a distinct structure and energy transfer pattern.

Isolation, properties and spatial site analysis of gamma subunits of B-phycoerythrin and R-phycoerythrin

Polysiphonia urceolata R-phycoerythrin and Porphyridium cruentum B-phycoerythrin were degraded with proteinaseK, and then the nearly native gamma subunits were isolated from the reaction mixture. The process of degradation of phycoerythrin with proteinaseK showed that the gamma subunit is located in the central cavity of (alpha beta)(6) hexamer of phycoerythrin. Comparative analysis of the spectra of the native phycoerythrin, the phycoerythrin at pH 12 and the isolated gamma subunit showed that the absorption peaks of phycoerythrobilins on alpha or beta subunit are at 535 nm (or 545 nm) and 565 nm, the fluorescence emission maximum at 580 nm; the absorption peak of phycoerythrobilins on the isolated gamma subunit is at 589 nm, the fluorescence emission peak at 620 nm which overlaps the absorption maximum of C-phycocyanin and perhaps contributes to the energy transfer with high efficiency between phycoerythrin and phycocyanin in phycobilisome; the absorption maximum of phycourobilin on the isolated gamma subunit is at 498 nm, which is the same as that in native phycoerythrin, and the fluorescence emission maximum at 575 nm.

Spectroscopic properties of the C-phycocyanin-allophycocyanin conjugate and the isolated phycobilisomes from Spirulina platensis

C-phycocyanin (CPC) and allophycocyanin (APC) were purified from Spirulina platensis, then the CPC was attached covalently to the APC by reacting their epsilon-amino groups. The excitation energy could be transferred from the CPC to the APC in the CPC-APC conjugate. Intact phycobilisomes (PBS), consisting of CPC, APC, colourless linker polypeptides, and APC B or L-cm, were isolated from S. platensis. Spectroscopic properties of the isolated PBSs kept at 20 degrees C for various times showed that the connection between the APC and the APC B or L-cm was looser than that between the CPC and the APC in the isolated PBSs. The CPC-APC conjugate was more stable than the isolated PBSs, and the linker polypeptides had a minor influence on the excitation energy transfer characteristic between different phycobiliproteins in the PBS.

Biosynthesis of fluorescent cyanobacterial allophycocyanin trimer in Escherichia coli

Allophycocyanin (APC), a cyanobacterial photosynthetic phycobiliprotein, functions in energy transfer as a light-harvesting protein. One of the prominent spectroscopic characteristics of APC is a strong red-shift in the absorption and emission maxima when monomers are assembled into a trimer. Previously, holo-APC alpha and beta subunits (holo-ApcA and ApcB) were successfully synthesized in Escherichia coli. In this study, both holo-subunits from Synechocystis sp. PCC 6803 were co-expressed in E. coli, and found to self-assemble into trimers. The recombinant APC trimer was purified by metal affinity and size-exclusion chromatography, and had a native structure identical to native APC, as determined by characteristic spectroscopic measurements, fluorescence quantum yield, tryptic digestion analysis, and molecular weight measurements. Combined with results from a study in which only the monomer was formed, our results indicate that bilin synthesis and the subsequent attachment to apo-subunits are important for the successful assembly of APC trimers. This is the first study to report on the assembly of recombinant ApcA and ApcB into a trimer with native structure. Our study provides a promising method for producing better fluorescent tags, as well as a method to facilitate the genetic analysis of APC trimer assembly and biological function.

Evolutionary analysis of phycobiliproteins: Implications for their structural and functional relationships

Phycobiliproteins, together with linker polypeptides and various chromophores, are basic building blocks of phycobilisomes, a supramolecular complex with a light-harvesting function in cyanobacteria and red algae. Previous studies suggest that the different types of phycobiliproteins and the linker polypeptides originated from the same ancestor. Here we retrieve the phycobilisome-related genes from the well-annotated and even unfinished cyanobacteria genomes and find that many sites with elevated d(N)/d(S) ratios in different phycobiliprotein lineages are located in the chromophore-binding domain and the helical hairpin domains (X and Y). Covariation analyses also reveal that these sites are significantly correlated, showing strong evidence of the functional-structural importance of interactions among these residues. The potential selective pressure driving the diversification of phycobiliproteins may be related to the phycobiliprotein-chromophore microenvironment formation and the subunits interaction. Sites and genes identified here would provide targets for further research on the structural-functional role of these residues and energy transfer through the chromophores.

电离层行进式扰动的GPS台网监测研究

In this dissertation, we investigated two types of traveling ionospheric disturbances (TIDs)/gravity waves (GWs) triggered separately by auroral energy input during super geomagnetic storms and solar terminator (ST) under quiet geomagnetic conditions (kp<3+) using TEC measurements from the global network of GPS receivers. Research into the generation and propagation of TIDs/GWs during storms greatly enhance our understandings on the evolution processes of energy transportation from the high-latitude’s magnetosphere to the low-latitude ionosphere and the conjugated effect of TIDs propagation between the northern and southern hemispheres. Our results revealed that the conjugacy of propagation direction between the northern and southern hemispheres was subject to the influence of Coriolis force. We also figure out the evolution processes of ionospheric disturbances at the global scale. These are important topics that had not been well addressed previously. In addition, we also obtained thee wave structures of medium scale TIDs excited by the solar terminator (ST) moving over the northern America and physical mechanisms involved. Our observations confirm that the ST is a stable and repetitive source of ionospheric wave disturbances and the evidence of solar terminator generated disturbances has been demonstrated experimentally via the GPS TEC measurement. The main researches and results of this dissertation are as follows. First, the global traveling ionospheric disturbances (TIDs) during the drastic magnetic storms of October 29–31, 2003 were analyzed using the Global Position System (GPS) total electron content (TEC) data observed in the Asian-Australian, European and North American sectors. We collected the most comprehensive set of the TEC data from more than 900 GPS stations on the International GNSS Services (IGS) website and introduce here a strategy that combines polynomial fitting and multi-channel maximum entropy spectral analysis to obtain TID parameters. Moreover, in collaboration with my thesis advisor, I have developed an imaging technique of 2-dimensional map of TIDs structures to obtain spatial and temporal maps of large scale traveling ionospheric disturbances (LSTIDs). The clear structures of TEC perturbations map during the passage of TIDs were displayed. The results of our study are summarized as follows: (1) Large-scale TIDs (LSTIDs) and medium-scale TIDs (MSTIDs) were detected in all three sectors after the sudden commencement (SC) of the magnetic storm, and their features showed longitudinal and latitudinal dependences. The duration of TIDs was longer at higher latitudes than at middle latitudes, with a maximum of about 16 h. The TEC variation amplitude of LSTIDs was larger in the North American sector than in the two other sectors. At the lower latitudes, the ionospheric perturbations were more complicated, and their duration and amplitude were relatively longer and larger. (2) The periods and phase speeds of TIDs were different in these three sectors. In Europe, the TIDs propagated southward; in North America and Asia, the TIDs propagated southwestward; in the near-equator region, the disturbances propagated with the azimuth (the angle of the propagation direction of the LSTIDs measured clockwise from due north with 0°) of 210° showing the influence of Coriolis force; in the Southern Hemisphere, the LSTIDs propagated conjugatedly northwestward. Both the southwestward and northeastward propagating LSTIDs are found in the equatorial region. These results mean that the Coriolis effect cannot be ignored for the wave propagation of LSTIDs and that the propagation direction is correlated with the polar magnetic activity. (3) The day (day of year: 301) before the SC (sudden commencement) of magnetic storm, we observed a sudden TEC skip disturbances (±10 TECU). It should be a response for the high flux of proton during the solar flare event, but not the magnetic storms. Next, the most comprehensive and dense GPS network’s data from North-America region were used in this paper to analyze the medium scale traveling ionospheric disturbances (MSTIDs) which were generated by the moving solar terminator during the quiet days in 2005. We applied the multi-channel maximum entropy spectral analysis to calculated TID parameters, and found that the occurrence of ST-MSTIDs depends on the seasonal variations. The results of our study are summarized as follows: (1) MSTIDs stimulated by the moving ST (ST-MSTIDs) are detected at mid-latitudes after the passage of the solar terminator with the life time of 2～3 hours and the variation amplitude of 0.2～0.8 TECU. Spectral analysis indicated that the horizontal wavelength, average period, horizontal phase velocity of the MSTIDs are around 300±150 km，150±80 m/s and 25±15 min, respectively. In addition, ST-MSTIDs have wave fronts elongating the moving ST direction and almost parallel to ST. (2) The statistical results demonstrate that the dusk MSTIDs stimulated by ST is more obvious than the dawn MSTIDs in summer. On the contrary, the more-pronounced dawn MSTIDs occurs in winter. (3) Further analysis indicates that the seasonal variations of ST-MSTIDs occurrence frequency are most probably related to the seasonal differences of the variations of EUV flux in the ionosphere region and recombination process during sunrise and sunset period at mid-latitudes. Statistical study of occurrence characteristics of TIDs using the GPS network in North-American and European during solar maximum, In conclusion, statistical studies of the propagation characteristics of TIDs, which excited by the two common origins including geomagnetic storms and moving solar terminator, were involved with global GPS TEC databasein this thesis. We employed the multichannel maximum entropy spectral analysis method to diagnose the characteristics of propagation and evolvement of ionospheric disturbances, also, the characteristics of their regional distribution and climatological variations were revealed by the statistic analysis. The results of these studies can improve our knowledge about the energy transfer in the solar-terrestrial system and the coupling process between upper and lower atmosphere (thermosphere-ionosphere-mesosphere). On the other hand, our results of the investigation on TIDs generated by particular linear origin such as ST are important for developing ionospheric irregularity physics and modeling the transionosphere radio wave propagation. Besides, the GPS TEC representation of the ST-generated ionospheric structure suggests a better possibility for investigating this phenomenon. Subsequently, there are scientific meaning of the result of this dissertation to deeply discuss the energy transfer and coupling in the ionosphere, as well as realistic value to space weather forecast in the ionosphere region.

Application of dynamical Lie algebraic method to atom-diatomic molecule scattering

The dynamical Lie algebraic approach developed by Alhassid and Levine combined with intermediate picture is applied to the study of translational-vibrational energy transfer in the collinear collision between an atom and an anharmonic oscillator. We find that the presence of the anharmonic terms indeed has an effect on the vibrational probabilities of the oscillator. The computed probabilities are in good agreement with those obtained using exact quantum method. It is shown that the approach of dynamical Lie algebra combining with intermediate picture is reasonable in the treating of atom-anharmonic oscillator scattering.