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A novel high light-inducible carotenoid-binding protein complex in the thylakoid membranes of Synechocystis PCC 6803.

Daddy S, Zhan J, Jantaro S, He C, He Q, Wang Q - Sci Rep (2015)

Bottom Line: We confirmed these findings by SEAD fluorescence cross-linking and anti-PsaD immuno-coprecipitation analyses.The expression of genes encoding the protein components of the HLCC was enhanced by high light illumination and artificial oxidative stress.Deletion of these proteins resulted in impaired state transition and increased sensitivity to oxidative and/or high light stress, as indicated by increased membrane peroxidation.

View Article: PubMed Central - PubMed

Affiliation: Department of Biology, University of Arkansas at Little Rock, Little Rock, AR 72204.

ABSTRACT
Synechocystis sp. PCC 6803 is a model cyanobacterium extensively used to study photosynthesis. Here we reveal a novel high light-inducible carotenoid-binding protein complex (HLCC) in the thylakoid membranes of Synechocystis PCC 6803 cells exposed to high intensity light. Zeaxanthin and myxoxanthophyll accounted for 29.8% and 54.8%, respectively, of the carotenoids bound to the complex. Using Blue-Native PAGE followed by 2D SDS-PAGE and mass spectrometry, we showed that the HLCC consisted of Slr1128, IsiA, PsaD, and HliA/B. We confirmed these findings by SEAD fluorescence cross-linking and anti-PsaD immuno-coprecipitation analyses. The expression of genes encoding the protein components of the HLCC was enhanced by high light illumination and artificial oxidative stress. Deletion of these proteins resulted in impaired state transition and increased sensitivity to oxidative and/or high light stress, as indicated by increased membrane peroxidation. Therefore, the HLCC protects thylakoid membranes from extensive photooxidative damage, likely via a mechanism involving state transition.

No MeSH data available.


Related in: MedlinePlus

Separation of the HLCC by Blue-Native PAGE followed by second dimension Tricine-SDS-PAGE.The HLCC fraction was carefully collected from the sucrose gradient and separated by BN PAGE (A), and the resulting single band was excised and denatured in 1.5 X SDS sample buffer and further separated on a 12–20% Tricine-SDS gel with 6 M urea (B). The proteins were visualized by silver staining and identified by mass spectrometry.
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f2: Separation of the HLCC by Blue-Native PAGE followed by second dimension Tricine-SDS-PAGE.The HLCC fraction was carefully collected from the sucrose gradient and separated by BN PAGE (A), and the resulting single band was excised and denatured in 1.5 X SDS sample buffer and further separated on a 12–20% Tricine-SDS gel with 6 M urea (B). The proteins were visualized by silver staining and identified by mass spectrometry.

Mentions: We further purified the fraction by Blue-Native PAGE. As shown in Fig. 2A, a single orange band was detected, indicating that there is one carotenoid-containing protein complex in the sucrose gradient fraction. We named this orange protein complex high light-inducible carotenoid-binding complex (HLCC). The orange Blue-Native band was then excised, denatured, and separated on a Tricine-SDS-PAGE gel (Fig. 2B), and four protein bands were revealed in the complex by silver staining. The bands were excised and identified by mass spectrometry. The three larger bands (from largest to smallest) contained the hypothetical protein Slr1128, the iron stress-induced chlorophyll-binding protein IsiA, and PsaD, a subunit of PSI, respectively. The fourth band at the lower end of the gel was identified as HliA and HliB.


A novel high light-inducible carotenoid-binding protein complex in the thylakoid membranes of Synechocystis PCC 6803.

Daddy S, Zhan J, Jantaro S, He C, He Q, Wang Q - Sci Rep (2015)

Separation of the HLCC by Blue-Native PAGE followed by second dimension Tricine-SDS-PAGE.The HLCC fraction was carefully collected from the sucrose gradient and separated by BN PAGE (A), and the resulting single band was excised and denatured in 1.5 X SDS sample buffer and further separated on a 12–20% Tricine-SDS gel with 6 M urea (B). The proteins were visualized by silver staining and identified by mass spectrometry.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4377637&req=5

f2: Separation of the HLCC by Blue-Native PAGE followed by second dimension Tricine-SDS-PAGE.The HLCC fraction was carefully collected from the sucrose gradient and separated by BN PAGE (A), and the resulting single band was excised and denatured in 1.5 X SDS sample buffer and further separated on a 12–20% Tricine-SDS gel with 6 M urea (B). The proteins were visualized by silver staining and identified by mass spectrometry.
Mentions: We further purified the fraction by Blue-Native PAGE. As shown in Fig. 2A, a single orange band was detected, indicating that there is one carotenoid-containing protein complex in the sucrose gradient fraction. We named this orange protein complex high light-inducible carotenoid-binding complex (HLCC). The orange Blue-Native band was then excised, denatured, and separated on a Tricine-SDS-PAGE gel (Fig. 2B), and four protein bands were revealed in the complex by silver staining. The bands were excised and identified by mass spectrometry. The three larger bands (from largest to smallest) contained the hypothetical protein Slr1128, the iron stress-induced chlorophyll-binding protein IsiA, and PsaD, a subunit of PSI, respectively. The fourth band at the lower end of the gel was identified as HliA and HliB.

Bottom Line: We confirmed these findings by SEAD fluorescence cross-linking and anti-PsaD immuno-coprecipitation analyses.The expression of genes encoding the protein components of the HLCC was enhanced by high light illumination and artificial oxidative stress.Deletion of these proteins resulted in impaired state transition and increased sensitivity to oxidative and/or high light stress, as indicated by increased membrane peroxidation.

View Article: PubMed Central - PubMed

Affiliation: Department of Biology, University of Arkansas at Little Rock, Little Rock, AR 72204.

ABSTRACT
Synechocystis sp. PCC 6803 is a model cyanobacterium extensively used to study photosynthesis. Here we reveal a novel high light-inducible carotenoid-binding protein complex (HLCC) in the thylakoid membranes of Synechocystis PCC 6803 cells exposed to high intensity light. Zeaxanthin and myxoxanthophyll accounted for 29.8% and 54.8%, respectively, of the carotenoids bound to the complex. Using Blue-Native PAGE followed by 2D SDS-PAGE and mass spectrometry, we showed that the HLCC consisted of Slr1128, IsiA, PsaD, and HliA/B. We confirmed these findings by SEAD fluorescence cross-linking and anti-PsaD immuno-coprecipitation analyses. The expression of genes encoding the protein components of the HLCC was enhanced by high light illumination and artificial oxidative stress. Deletion of these proteins resulted in impaired state transition and increased sensitivity to oxidative and/or high light stress, as indicated by increased membrane peroxidation. Therefore, the HLCC protects thylakoid membranes from extensive photooxidative damage, likely via a mechanism involving state transition.

No MeSH data available.


Related in: MedlinePlus