Excitation energy relaxation in a symbiotic cyanobacterium, Prochloron didemni, occurring in coral-reef ascidians, and in a free-living cyanobacterium, Prochlorothrix hollandica.

The marine cyanobacterium Prochloron is a unique photosynthetic organism that lives in obligate symbiosis with colonial ascidians. We compared Prochloron harbored in four different host species and cultured Prochlorothrix by means of spectroscopic measurements, including time-resolved fluorescence, to investigate host-induced differences in light-harvesting strategies between the cyanobacteria. The light-harvesting efficiency of photosystems including antenna Pcb, PS II-PS I connection, and pigment status, especially that of PS I Red Chls, were different among the four samples. We also discuss relationships between these observed characteristics and the light conditions, to which Prochloron cells are exposed, influenced by distribution pattern in the host colonies, presence or absence of tunic spicules, and microenvironments within the ascidians' habitat.

Localization of Pcb antenna complexes in the photosynthetic prokaryote Prochlorothrix hollandica.

The freshwater filamentous green oxyphotobacterium Prochlorothrix hollandica is an unusual oxygenic photoautotrophic cyanobacterium differing from most of the others by the presence of light-harvesting Pcb antenna binding both chlorophylls a and b and by the absence of phycobilins. The pigment-protein complexes of P. hollandica SAG 10.89 (CCAP 1490/1) were isolated from dodecylmaltoside solubilized thylakoid membranes on sucrose density gradient and characterized by biochemical, spectroscopic and immunoblotting methods. The Pcb antennae production is suppressed by high light conditions (>200 mumol photons m(-2) s(-1)) in P. hollandica. PcbC protein was found either in higher oligomeric states or coupled to PS I (forming antenna rings around PS I). PcbA and PcbB are most probably only very loosely bound to photosystems; we assume that these pigment-protein complexes function as low light-induced mobile antennae. Further, we have detected alpha-carotene in substantial quantities in P. hollandica thylakoid membranes, indicating the presence of chloroplast-like carotenoid synthetic pathway which is not present in common cyanobacteria.

Salinity tolerance of the chlorophyll b-synthesizing cyanobacterium Prochlorothrix hollandica strain SAG 10.89.

Ecophysiological investigations on the salinity acclimation of the cyanobacterium Prochlorothrix hollandica SAG 10.89 led to significantly revised salinity tolerance limits. Besides potential effects of cultivation techniques, clear ion composition effects mainly explain formerly described hypersensitivity to NaCl-mediated salinity and lack of osmolyte detection. An extraordinarily broad plasticity of cellular chlorophyll a/b ratios occurred with variations of NaCl-induced salinity. Photosynthesis characteristics, pigment regulation, respiration, and biomass yield in growth medium with field-like ion composition indicated generally reduced acclimation pressure. A simultaneously significant increase in osmolyte (sucrose) accumulation indicated more efficient osmotic acclimation. Minor growth inhibition up to salinities of 10 practical salinity units enlarged the potential habitat of P. hollandica but at the most to about 300,000 km2 in the Baltic Sea. This supports probable observations of Prochlorothrix sp. in phytoplankton assemblages of open waters in Baltic Sea-monitoring studies. Brackish habitats differ from so far known habitats of Prochlorothrix spp. in turbidity, productivity, and plankton composition. Adjusted physiological features dispel fundamental doubts on the establishment of filamentous prochlorophytes in brackish waters.

Association of chlorophyll a/b-binding Pcb proteins with photosystems I and II in Prochlorothrix hollandica.

Action spectra for photosystem II (PSII)-driven oxygen evolution and of photosystem I (PSI)-mediated H(2) photoproduction and photoinhibition of respiration were used to determine the participation of chlorophyll (Chl) a/b-binding Pcb proteins in the functions of pigment apparatus of Prochlorothrix hollandica. Comparison of the in situ action spectra with absorption spectra of PSII and PSI complexes isolated from the cyanobacterium Synechocystis 6803 revealed a shoulder at 650 nm that indicated presence of Chl b in the both photosystems of P. hollandica. Fitting of two action spectra to absorption spectrum of the cells showed a chlorophyll ratio of 4:1 in favor of PSI. Effective antenna sizes estimated from photochemical cross-sections of the relevant photoreactions were found to be 192+/-28 and 139+/-15 chlorophyll molecules for the competent PSI and PSII reaction centers, respectively. The value for PSI is in a quite good agreement with previous electron microscopy data for isolated Pcb-PSI supercomplexes from P. hollandica that show a trimeric PSI core surrounded by a ring of 18 Pcb subunits. The antenna size of PSII implies that the PSII core dimers are associated with approximately 14 Pcb light-harvesting proteins, and form the largest known Pcb-PSII supercomplexes.

Characteristic fragmentation of bacteriohopanepolyols during atmospheric pressure chemical ionisation liquid chromatography/ion trap mass spectrometry.

Bacteriohopanepolyols (BHPs) fragment via characteristic pathways during atmospheric pressure chemical ionisation liquid chromatography/ion trap mass spectrometry (APCI-LC/MS(n)). Comparison of the MS(2) spectra of bacteriohopane-32,33,34,35-tetrol (BHT) and 2 beta-methylbacteriohopane-32,33,34,35-tetrol has confirmed the previously proposed ring-C cleavage occurring between C-9 and 11 and C-8 and 14. This fragmentation, diagnostic of all hopanoids, also occurs in BHPs containing an amino group (-NH(2)) at C-35 although the higher relative stability of the ion limits this fragmentation to a minor process after protonation of the basic nitrogen function. Studies of a number of cell cultures including a prochlorophyte (Prochlorothrix hollandica) and a cyanobacterium (Chlorogloeopsis LA) demonstrate the power of this technique to detect composite BHPs with a complex biological functionality at C-35. We also report the first observation of intact pentafunctionalised bacteriohopanepolyols using this method.

Mutagenesis of prochlorothrix plastocyanin reveals additional features in photosystem I interactions.

Three surface residues of plastocyanin from Prochlorothrix hollandica have been modified by site-directed mutagenesis. Changes have been made in methionine 33, located in the hydrophobic patch of the copper protein, and in arginine 86 and proline 53, both located in the eastern hydrophilic area. The reactivity toward photosystem I of single mutants M33N, P53A, P53E, R86Q, R86E, and the double mutant M33N/P14L has been studied by laser flash absorption spectroscopy. All the mutations yield increased reactivity of plastocyanin toward photosystem I as compared with wild type plastocyanin, thus indicating that in Prochlorothrix electron donation to photosystem I is not optimized. The most drastic increases in the intracomplex electron transfer rate are obtained with mutants in methionine 33, whereas replacing arginine 86 only modestly affects the plastocyanin-photosystem I equilibrium constant for complex formation. Mutations at position 53 also promote major changes in the association of plastocyanin with photosystem I, yielding a change from a mechanism involving complex formation to a simpler collisional interaction. Molecular dynamics calculations indicate that mutations at position 33 promote changes in the H-bond network around the copper center. The comparative kinetic analysis of the reactivity of Prochlorothrix plastocyanin mutants toward photosystem I from other cyanobacteria reveals that mutations M33N, P53A, and P53E result in enhanced general reactivity.

The unique proline of the Prochlorothrix hollandica plastocyanin hydrophobic patch impairs electron transfer to photosystem I.

A number of surface residues of plastocyanin from Prochlorothrix hollandica have been modified by site-directed mutagenesis. Changes have been made in amino acids located in the amino-terminal hydrophobic patch of the copper protein, which presents a variant structure as compared with other plastocyanins. The single mutants Y12G, Y12F, Y12W, P14L, and double mutant Y12G/P14L have been produced. Their reactivity toward photosystem I has been analyzed by laser flash absorption spectroscopy. Plots of the observed rate constant with all mutants versus plastocyanin concentration show a saturation profile similar to that with wild-type plastocyanin, thus suggesting the formation of a plastocyanin-photosystem I transient complex. The mutations do not induce relevant changes in the equilibrium constant for complex formation but induce significant variations in the electron transfer rate constant, mainly with the two mutants at proline 14. Additionally, molecular dynamics calculations indicate that mutations at position 14 yield small changes in the geometry of the copper center. The comparative kinetic analysis of the reactivity of plastocyanin mutants toward photosystem I from different organisms (plants and cyanobacteria) reveals that reversion of the unique proline of Prochlorothrix plastocyanin to the conserved leucine of all other plastocyanins at this position enhances the reactivity of the Prochlorothrix protein.

Sequence and functional characterization of RNase P RNA from the chl alb containing cyanobacterium Prochlorothrix hollandica.

Only a few complete sequences and very limited functional data are available for the catalytic RNA component of cyanobacterial RNase P. The RNase P RNA from the chl alb containing cyanobacterium Prochlorothrix hollandica belongs to a rarely found structural subtype with an extended P15/16 domain. We have established conditions for optimal in vitro ribozyme activity, and determined the kinetic parameters for cleavage of pre-tRNA(Tyr). Analysis of pre-tRNA mutants revealed that the T-stem sequence only plays a modulating role, whereas the CCA end is essential for efficient product formation.