pH‐dependent protonation of surface carboxylate groups in PsbO enables local buffering and triggers structural changes

Photosystem II (PSII) catalyzes the splitting of water, releasing protons and dioxygen. Its highly conserved subunit PsbO extends from the oxygen‐evolving center (OEC) into the thylakoid lumen and stabilizes the catalytic Mn4CaO5 cluster. The high degree of conservation of accessible negatively charged surface residues in PsbO suggests additional functions, as local pH buffer or by affecting the flow of protons. For this discussion, we provide an experimental basis, through the determination of pKa values of water‐accessible aspartate and glutamate side‐chain carboxylate groups by means of NMR. Their distribution is strikingly uneven, with high pKa values around 4.9 clustered on the luminal PsbO side and values below 3.5 on the side facing PSII. pH‐dependent changes in backbone chemical shifts in the area of the lumen‐exposed loops are observed, indicating conformational changes. In conclusion, we present a site‐specific analysis of carboxylate group proton affinities in PsbO, providing a basis for further understanding of proton transport in photosynthesis

A review of the scientific knowledge of the seascape off Dronning Maud Land, Antarctica

Despite the exclusion of the Southern Ocean from assessments of progress towards achieving the Convention on Biological Diversity (CBD) Strategic Plan, the Commission for the Conservation of Antarctic Marine Living Resources (CCAMLR) has taken on the mantle of progressing efforts to achieve it. Within the CBD, Aichi Target 11 represents an agreed commitment to protect 10% of the global coastal and marine environment. Adopting an ethos of presenting the best available scientific evidence to support policy makers, CCAMLR has progressed this by designating two Marine Protected Areas in the Southern Ocean, with three others under consideration. The region of Antarctica known as Dronning Maud Land (DML; 20 degrees W to 40 degrees E) and the Atlantic sector of the Southern Ocean that abuts it conveniently spans one region under consideration for spatial protection. To facilitate both an open and transparent process to provide the vest available scientific evidence for policy makers to formulate management options, we review the body of physical, geochemical and biological knowledge of the marine environment of this region. The level of scientific knowledge throughout the seascape abutting DML is polarized, with a clear lack of data in its eastern part which is presumably related to differing levels of research effort dedicated by national Antarctic programmes in the region. The lack of basic data on fundamental aspects of the physical, geological and biological nature of eastern DML make predictions of future trends difficult to impossible, with implications for the provision of management advice including spatial management. Finally, by highlighting key knowledge gaps across the scientific disciplines our review also serves to provide guidance to future research across this important region.Peer reviewe

A review of the scientific knowledge of the seascape off Dronning Maud Land, Antarctica

Despite the exclusion of the Southern Ocean from assessments of progress towards achieving the Convention on Biological Diversity (CBD) Strategic Plan, the Commission for the Conservation of Antarctic Marine Living Resources (CCAMLR) has taken on the mantle of progressing efforts to achieve it. Within the CBD, Aichi Target 11 represents an agreed commitment to protect 10% of the global coastal and marine environment. Adopting an ethos of presenting the best available scientific evidence to support policy makers, CCAMLR has progressed this by designating two Marine Protected Areas in the Southern Ocean, with three others under consideration. The region of Antarctica known as Dronning Maud Land (DML; 20°W to 40°E) and the Atlantic sector of the Southern Ocean that abuts it conveniently spans one region under consideration for spatial protection. To facilitate both an open and transparent process to provide the vest available scientific evidence for policy makers to formulate management options, we review the body of physical, geochemical and biological knowledge of the marine environment of this region. The level of scientific knowledge throughout the seascape abutting DML is polarized, with a clear lack of data in its eastern part which is presumably related to differing levels of research effort dedicated by national Antarctic programmes in the region. The lack of basic data on fundamental aspects of the physical, geological and biological nature of eastern DML make predictions of future trends difficult to impossible, with implications for the provision of management advice including spatial management. Finally, by highlighting key knowledge gaps across the scientific disciplines our review also serves to provide guidance to future research across this important region.publishedVersio

First results from the L3+C experiment at CERN

The L3+C experiment combines the high-precision spectrometer of the L3 detector at LEP, CERN, with a small air shower array. The momenta of cosmic ray induced muons can be measured from 20 to 2000 GeV/c. During the 1999 data taking period 5 billion muon events were recorded in the spectrometer. From April until mid Summer 2000 an additional 3 billion muon events have been recorded as well as 25 million air shower events. Here the first results on the muon momentum spectrum and charge ratio will be presented

Untersuchung von protonierungsdynamischen Prozessen in lichtabsorbierenden und umwandelnden Proteinen mittels NMR

A way to induce structural changes in proteins is through alterations of the protonation state, a principle which is referred to as ‘protonation dynamics’. The two proteins investigated in this work, PsbO and Cph1, are both involved in processes attributed to protonation dynamics, and are studied by solution and solid-state NMR. Photosynthesis produces highly reactive compounds, whose concentrations must be strictly monitored and regulated. At the oxygen evolving centre (OEC) of photosystem II (PSII) - a multiprotein complex spanning the thylakoid membrane - the water-splitting reaction takes place and generates a high proton concentration at the proton release site. PsbO, being in close proximity to the OEC and bearing a significant number of surface carboxyl groups, is likely involved in proton storage or translocation to the lumen. This work investigates the protonation state of the surface aspartic and glutamic acid side chain carboxyl groups of the beta-barrel of PsbO to provide information about a possible proton transfer pathway. Solution NMR experiments were employed. pKa values of 21 carboxylate groups, of which 9 are aspartate and 12 glutamate residues, were determined by the application of 2D titration experiments. The determined pKa values were plotted onto the crystal structure, resulting in an uneven distribution with values below 3.5 clustered on the side of the protein facing PSII whereas values around 4.9 were located at the luminal side of PsbO. This study shows that the bilateral surface pKa distribution enables PsbO to serve as a luminal buffer and potential acceptor of protons, generated during PSII activity. Phytochromes are red and far-red light sensing photoreceptors, present in plants, fungi, and cyanobacteria that control developmental processes such as seed germination and flowering. Sedimentation NMR (sedNMR) and dynamic nuclear polarization (DNP) were combined at high field (800 MHz) to study the so-called ‘tongue’ region of the cyanobacterial phytochrome Cph1∆2, a structural element conserved in all phytochromes. This element is supposed to change its secondary structure from beta-sheet to alpha-helix upon light irradiation. We want to observe the transition in one protein by studying and comparing Cph1∆2 in the Pr and Pfr state. The application of DNP at high fields is not trivial, since enhancements decline and standard measurement conditions can lead to partial unfolding of the proteins investigated. For this reason, new buffer conditions were tested and two polarizing agents, bctol-M and M-TinyPol, were characterised. The performance of bcTol-M was convincing, showing a similar enhancement but higher stability than M-TinyPol. The combination of a selective amino acid labelling scheme and pulse sequence filtering led to reduced spectral overlap, enabling the assignment of resonances within the 58 kDa cyanobacterial phytochrome Cph1∆2 at DNP measurement conditions. By the comparison of 3D hCaNCOCa spectra, recorded on Pr and Pfr state Cph1∆2, a strong indication for structural rearrangements within the ‘tongue’ region was found.Strukturänderungen können in einem Protein durch die Veränderung des Protonierungszustandes hervorgerufen werden, ein Prinzip das unter den Begriff ‚Protonierungsdynamik‘ fällt. Die zwei in dieser Arbeit untersuchten Proteine, PsbO und Cph1, sind beide an Prozessen beteiligt, die der Protonierungsdynamik zugeschrieben werden. Sie werden mittels Lösungs- und Festkörper-NMR untersucht. Bei der Photosynthese entstehen hochreaktive Verbindungen, deren Konzentrationen streng überwacht und reguliert werden müssen. Am sauerstoffproduzierenden Komplex (OEC) des Photosystems II (PSII) – einem Multiproteinkomplex, der die Thylakoidmembran durchzieht – findet die Wasserspaltung statt. Das führt zu einer erhöhten Protonenkonzentration am Ort der Protonenfreisetzung. PsbO befindet sich in unmittelbarer Nähe des OEC und trägt eine hohe Anzahl an Oberflächencarboxylgruppen. Es ist wahrscheinlich, dass PsbO als Protonenspeicher dient oder an dem Transport der Protonen ins Thylakoidlumen beteiligt ist. In dieser Arbeit wurde der Protonierungszustand der Asparaginsäure- und Glutaminsäure-Seitenkettencarboxylgruppen von PsbO mit Lösungs-NMR untersucht um Informationen über einen möglichen Protonentransferweg zu erhalten. pKs – Werte von 21 Carboxylat- gruppen, darunter 9 Aspartat- und 12 Glutamatreste, konnten durch 2D- Titrationsexperimente bestimmt werden. Die ermittelten pKs – Werte wurden auf die Kristallstruktur von PsbO übertragen, wobei eine ungleichmäßige Verteilung beobachtet wurde. Auf der, dem PSII zugewandten Seite des Proteins, nehmen die Carboxylatgruppen pKs – Werte unter 3,5 an, wohingegen auf der luminalen Seite pKs – Werte um 4,9 beobachtet wurden. Mit dieser Studie konnte gezeigt werden, dass die Verteilung von pKs – Werten auf der Oberfläche von PsbO dazu führt, dass das Protein als luminaler Puffer fungieren kann und ein potenzieller Akzeptor von Protonen ist, die während der PSII-Aktivität entstehen. Phytochrome sind Photorezeptoren, die Licht aus dem roten und infraroten Bereich wahrnehmen. Sie kommen in Pflanzen, Pilzen und Cyanobakterien vor und steuern Entwicklungsprozesse wie Samenkeimung und Blüte. In dieser Arbeit wurden Sedimentations-NMR und dynamische Kernspinpolarisation bei hohen Magnetfeldstärken (800 MHz) kombiniert um die sogenannte ‚Zungen‘-Region des cyanobakteriellen Phytochroms Cph1∆2 zu untersuchen, ein Strukturelement welches in allen Phytochromen konserviert ist. Es wird angenommen, dass dieses Element bei Lichteinstrahlung seine Sekundärstruktur von einem β-Faltblatt zu einer α-Helix ändert. In dieser Arbeit soll die Strukturänderung in einem einzigen Protein beobachtet werden, indem Cph1∆2 im Pfr und Pr Zustand untersucht und verglichen wird. Der Einsatz von DNP bei hohen Feldstärken ist nicht trivial, da die Signalverstärkung im Vergleich zu geringeren Magnetfeldern reduziert ist, außerdem kommt es unter den Standardmessbedingungen zur teilweisen Entfaltung der zu untersuchenden Proteine. Aus diesem Grund wurden neue Pufferbedingungen getestet und zwei Radikale, bcTol-M und M-TinyPol, charakterisiert und verglichen. bcTol zeigte eine ähnliche Signalverstärkung wie M-TinyPol, aber eine höhere Stabilität. Durch die Kombination eines spezifischen Aminosäuremarkierungsmusters mit einer selektiven Pulssequenz konnte die Überlappung von Signalen verringert werden. Dadurch wurde die Zuordnung von Resonanzen des 58 kDa cyanobakteriellen Phytochroms Cph1∆2 unter DNP Messbedingungen ermöglicht. Der Vergleich von 3D hCaNCOCa Spektren, aufgenommen von Cph1∆2 im Pr und Pfr Zustand, deutete auf strukturelle Umlagerungen im Bereich der ‚Zungen‘-Region hin