Quadrupol Massenspektrometer mit elektrostatischer Ionenfalle als Detektor und Nano-Flüssigkeitschromatographie System
Final Report Abstract
The laboratory is interested in understanding the regulation of photosynthesis. Acclimation towards excess light requires a mechanism designated as non-photochemical quenching (NPQ). In Chla-mydomonas reinhardtii, qE, the fastest component of NPQ, is mainly facilitated by LHCSR3. The new mass spectrometer permitted quantitative proteome analyses allowing insights into LHCSR3 function and regulation. Our new data indicated that the PSBR subunit of PSII is required for efficient binding of LHCSR3 to PSII-LHCII supercomplexes, suggesting that LHCSR3 is in close contact to the minor antenna proteins LHCB4 and LHCB5. Moreover, the N-terminus of LHCSR3 was phosphorylated in an STT7-dependent manner, while the C-terminal phosphorylation of LHCSR3 is STT7 independent (Bergner et al, 2015). The new mass spectrometer also allowed very sensitive quantitation of protein phosphorylation, which was used to quantity amounts of phosphory-lated LHCSR3 in a time course experiment. In another project line, we are interested in calcium (Ca2+) dependent regulation of photosynthesis. Ca2+ and redox signaling play important roles in acclimation processes from archaea to eukaryotic organisms. Herein we characterized a unique protein from Chlamydomonas reinhardtii that has the competence to integrate Ca2+ and redox-related signaling. This protein, originally identified in a proteomic study, was designated as calredoxin (CRX), as it combines four Ca2+-binding EF-hands and a thioredoxin (TRX) domain. A protein crystal structure of CRX, at 1.6 Å resolution, revealed an unusual calmodulin-fold of the Ca2+-binding EF-hands, which is functionally linked via an inter-domain communication path with the enzymatically active TRX domain. CRX is chloroplast-localized and interacted with a chloroplast 2-Cys-peroxiredoxin (PRX1). Ca2+-binding to CRX is critical for its TRX activity and for efficient binding and reduction of PRX1. Thereby CRX represents a new class of Ca2+-dependent "sensor-responder" proteins. Genetically engineered Chlamydomonas strains with strongly diminished amounts of CRX, revealed altered photosynthetic electron transfer, a decreased expression of thioredoxin f upon high light treatment and impact in ROS defense underpinning a function of CRX in chloroplast redox and stress acclimation. Quantitative proteomics data revealing a role of CRX in ROS defence were generated with the novel mass spectrometer. Moreover, we employed mass spectrometry to analyze temperature-induced remodeling of the photosynthetic machinery in the thermophilic alga Cyanidioschyzon merolae. The new mass spectrometer enabled us shed light on the N-glycosylation pathway of the green colonial alga Botryococcus braunii. Moreover, we are using mass spectrometry to elucidate the N-glycosylation pathway of Chlamydomonas reinhardtii.
Publications
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(2015) Photosystem II Subunit R Is Required for Efficient Binding of Light-Harvesting Complex Stress-Related Protein3 to Photosystem II-Light-Harvesting Supercomplexes in Chlamydomonas reinhardtii. Plant physiology 167: 1566-1578
Xue H, Tokutsu R, Bergner SV, Scholz M, Minagawa J, Hippler M
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(2015) STATE TRAN-SITION7-Dependent Phosphorylation Is Modulated by Changing Environmental Conditions, and Its Absence Triggers Remodeling of Photosynthetic Protein Complexes. Plant physiology 168: 615-634
Bergner SV, Scholz M, Trompelt K, Barth J, Gabelein P, Steinbeck J, Xue H, Clowez S, Fucile G, Goldschmidt- Clermont M, Fufezan C, Hippler M
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(2016) Calredoxin represents a novel type of calcium-dependent sensor-responder connected to redox regulation in the chloroplast. Nat Commun 7: 11847
Hochmal AK, Zinzius K, Charoenwattanasatien R, Gabelein P, Mutoh R, Tanaka H, Schulze S, Liu G, Scholz M, Nordhues A, Offenborn JN, Petroutsos D, Finazzi G, Fufezan C, Huang K, Kurisu G, Hippler M
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(2016) Identification of Haloferax volcanii Pilin N- Glycans with Diverse Roles in Pilus Biosynthesis, Adhesion, and Microcolony Formation. The Journal of biological chemistry 291: 10602-10614
Esquivel RN, Schulze S, Xu R, Hippler M, Pohlschroder M
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(2017) ArtA-Dependent Processing of a Tat Substrate Containing a Conserved Tripartite Structure That Is Not Localized at the C Terminus. Journal of bacteriology 199
Abdul Halim MF, Stoltzfus JD, Schulze S, Hippler M, Pohlschroder M
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(2017) Comparative transcriptome and proteome analysis reveals a global impact of the nitrogen regulators AreA and AreB on secondary metabolism in Fusarium fujikuroi. PloS one 12: e0176194
Pfannmuller A, Leufken J, Studt L, Michielse CB, Sieber CMK, Guldener U, Hawat S, Hippler M, Fufezan C, Tudzynski B
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(2017) Conservation of core complex subunits shaped the structure and function of photosystem I in the secondary endosymbiont alga Nannochloropsis gaditana. The New phytologist 213: 714-726
Alboresi A, Le Quiniou C, Yadav SK, Scholz M, Meneghesso A, Gerotto C, Simionato D, Hippler M, Boekema EJ, Croce R, Morosinotto T
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(2017) Identification of methylated GnTI-dependent N-glycans in Botryococcus brauni. The New phytologist 215: 1361-1369
Schulze S, Urzica E, Reijnders M, van de Geest H, Warris S, Bakker LV, Fufezan C, Martins Dos Santos VAP, Schaap PJ, Peters SA, Hippler M
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(2017) pyQms enables universal and accurate quantification of mass spectrometry data. Molecular & cellular proteomics : MCP
Leufken J, Niehues A, Sarin P, Wessel F, Hippler M, Leidel SA, Fufezan C
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(2017) Temperature-Induced Remodeling of the Photosynthetic Machinery Tunes Photosynthesis in the Thermophilic Alga Cyanidioschyzon merolae. Plant physiology 174: 35-46
Nikolova D, Weber D, Scholz M, Bald T, Scharsack JP, Hippler M
