Dynamics in stress-regulated betaine transport and role of pathogen-relevant choline transport

Transport processes across the membrane are essential to ensure survival of every living cell. Therefore, the exchange of membrane impermeable molecules is mediated by specific transport proteins, which are embedded in the lipid bilayer. One important class comprises secondary active transporters, which couple very efficiently the uphill transport of the main substrate against its concentration gradient to the downhill transport of an additional substrate. These transporters are widely distributed among all kingdoms of life and accomplish many crucial functions. One function is to counteract the deleterious effect of hyperosmotic stress in bacteria. Several members of the BCCT (betaine-choline-carnitinetransport) family of secondary transporters mediate osmostress protection by the accumulation of the compatible solute betaine or its precursor choline (Lamark et al., 1991; Peter et al., 1996; Ziegler et al., 2010). Besides osmo-dependent sodium or proton-coupled symporters, the BCCT family includes few rare representatives of osmo-independent transporters such as the substrate:product antiporter CaiT from E. coli (Jung et al., 2002; Ziegler et al., 2010). The best-characterized member of the BCCT family is the sodium-coupled betaine transporter BetP from Corynebacterium glutamicum. BetP together with the ABCtransporter OpuA and the H+-solute symporter ProP, became a paradigm for osmoregulated osmolyte transport. Although, all three transporters were extensively studied, the general mechanism of osmoregulation is still far from being understood. Thus, one task of this thesis was to elucidate further the regulatory properties of BetP. BetP is tightly regulated by osmotic stress and is able to increase its basal betaine uptake activity dramatically upon elevated osmolalities within one second (Peter et al., 1998a). The osmotic stress is sensed by BetP via two stimuli, one is the increase of the internal K+ concentration above a threshold of 220 mM (Rübenhagen et al., 2001), the second is related to a change in the physical state of the membrane (Maximov et al., 2014). So far, several solved crystal structures in combination with functional and computational analysis provided insights into the coupling mechanism of betaine and its co-substrate sodium (Khafizov et al., 2012; Perez et al., 2012). Despite the wealth of data, the precise regulatory mechanism of trimeric BetP is still unclear.Transportprozesse durch die Membran sind essentiell um das Überleben jeder lebenden Zelle zu gewährleisten. Daher wird der Austausch Membranundurchlässiger Moleküle durch spezifische Transportproteine, die innerhalb der Lipid-Doppelschicht eingebettet vorliegen, ermöglicht. Eine wichtige Klasse dabei sind die sekundär aktiven Transporter, die den Transport des ersten Substrates entgegen das Konzentrationsgefälle sehr effizient an den Transport eines weiteren Substrates in Richtung Konzentrationsgefälle koppeln. Diese Transporter sind in allen Reichen des Lebens weit verbreitet, da sie äußerst wichtige Funktionen ausführen. Eine Funktion ist es, dem schädlichen Effekt von hyperosmotischen Stress in Bakterien entgegenzuwirken. Mehrere Mitglieder der BCCT (Betain-Cholin-Carnitin-Transport)-Familie von sekundären Transportern schützen vor osmotischem Stress durch die Akkumulation von dem kompatiblem Solut Betain oder dessen Vorstufe, dem Cholin (Lamark et al., 1991; Peter et al., 1996; Ziegler et al., 2010). Abgesehen von osmotisch abhängigen Natrium- und Proton-gekoppelten Symportern, gehören auch einige wenige osmotisch unabhängige Transporter, wie der Substrat:Endprodukt Antiporter CaiT von E. coli, zu der BCCT-Familie (Jung et al., 2002; Ziegler et al., 2010). Das bestcharakterisierte Mitglied der BCCT-Familie ist der Natrium-gekoppelte Betain-Transporter BetP von Corynebacterium glutamicum. BetP zählt zusammen mit dem ABC-Transporter OpuA und dem H+-Solut Symporter ProP als Paradigma für osmotisch regulierten Osmolyt-Transport. Obwohl alle drei Transporter ausgiebig untersucht worden sind, ist der generelle Mechanismus der Osmoregulation noch weitgehend unverstanden. Demnach war eine Aufgabe, die Aufklärung weiterer regulatorischen Eigenschaften von BetP

Mozliwosci wykorzystania szklistych nawozow mineralnych w produkcji ogrodniczej

Opracowano szklą z układu K₂O-MgO-CaO-P₂O₅-SiO₂, które mogą być użyte jako ekologiczne nawozy sztuczne o wydłużonym działaniu. Szkła te są trudno rozpuszczalne w wodzie. Dobrze rozpuszczają się natomiast w 2 % roztworze kwasu cytrynowego, co wskazuje, że zawarty w nim potas, fosfor i magnez może być przyswajany przez rośliny. Szkła o odpowiednio wzbogaconym składzie stają się również źródłem Mo, Zn, B, Mn, Fe, Cu i innych mikroelementów. Rozpuszczalność szkieł reguluje się ich składem. Szkła wolniej rozpuszczalne w formie wełny mineralnej, granul, płatków proponuje się wykorzystać jako zasobne, aktywne podłoża ogrodnicze lub ich składniki.K₂O-MgO-CaO-P₂O₅-SiO₂ glasses were obtained by melting of apatite, serpentinite and K₂CO₃ mixtures. Microelements can be also introduced to the composition of glass. Solubility of the glasses in 2 % citric acid solution was studied and relation between their chemical composition and solubility determined. This increases as the P₂O₅ and K₂O content in glass increase. Glasses containing 10 - 15 % P₂O₅ and more than 5 % K₂O and set of microelements (0.4 - 5 %) can be used as ecological, controlled release fertilizers. Application of less soluble glasses, containing about 5 % P₂O₅, 5 % K₂O and microelements, as artificial horticultural soils is proposed

Identification of an osmo-dependent and an osmo-independent choline transporter in Acinetobacter baylyi: implications in osmostress protection and metabolic adaptation

Members of the genus Acinetobacter are well known for their metabolic versatility that allows them to adapt to different ecological niches. In previous studies, we have demonstrated that Acinetobacter baylyi ADP1 can cope with high salinities by uptake and accumulation of the well-known compatible solute glycine betaine. Here, we demonstrate that addition of choline restores growth at high salinities. We further show that choline was actively taken up by the cells and converted to glycine betaine. Uptake of choline was induced by high salinity and the presence of choline in the growth medium. At high salinities, glycine betaine was accumulated in the cells whereas in the absence of osmotic stress it was exported. Inspection of the genome sequence followed by mutant studies led to the identification of two genes encoding secondary transporters (BetT1 and BetT2) of the betaine-choline-carnitine transporter (BCCT) family. The BetT1 transporter lacks an extended C-terminal domain usually found in osmoregulated choline BCCTs. BetT1 was found to facilitate osmolarity-independent choline transport most likely by a uniport mechanism. We propose that BetT1 does not primarily function in osmoadaptation but might play a role in metabolic adaptation to choline-rich environments