Subcellular protein localization (cell envelope) in Phaeobacter inhibens DSM 17395

Phaeobacter inhibens DSM 17395 is a metabolically versatile, secondary metabolite producing and surface colonizing member of the alphaproteobacterial Roseobacter clade. Proteins compartmentalized across the Gram-negative cell envelope are expected to be relevant for the habitat success of P. inhibens DSM 17395. Subcellular fractionation was followed by gel- or nano-LCbased separation of proteins and peptides, respectively. Subsequent MS-based identification of in total 1187 proteins allowed allocation to cytoplasm (303 proteins), cytoplasmic membrane (346), periplasm (325), outer membrane (76), and extracellular milieu (22). Multidimensional scaling was used to visualize the spreading of heuristically allocated proteins across the five different compartments. Experimentally inferred subcellular protein localization was compared with PSORTb prediction of protein secretion and membrane localization. Determined subcellular localizations of identified proteins were interpreted to reconstruct the functional traits of the different cell envelope compartments, in particular protein secretion and sorting, direct effector molecule transit, and cell envelope biogenesis. From a proteogenomic perspective, functional prediction of 74 genes (including 17 coding for proteins of hitherto unknown function) could be refined

Einfluß von VEGF auf die Reendothelialisierung nach mikrochirurgischer Gefäßanastomose

Charles Heal's Ark - A91 - photographed 26 May 1934. Digitisation and record funded by the Pilgrim Trust

Pathways and substrate-specific regulation of amino acid degradation in Phaeobacter inhibens DSM 17395 (archetype of the marine Roseobacter clade)

Combining omics and enzymatic approaches, catabolic routes of nine selected amino acids (tryptophan, phenylalanine, methionine, leucine, isoleucine, valine, histidine, lysine and threonine) were elucidated in substrate-adapted cells of Phaeobacter inhibensDSM 17395 (displaying conspicuous morphotypes). The catabolic network [excluding tricarboxylic acid (TCA) cycle] was reconstructed from 71 genes (scattered across the chromosome; one-third newly assigned), with 69 encoded proteins and 20 specific metabolites identified, and activities of 10 different enzymes determined. For example, Ph.inhibensDSM 17395 does not degrade lysine via the widespread saccharopine pathway but might rather employ two parallel pathways via 5-aminopentanoate or 2-aminoadipate. Tryptophan degradation proceeds via kynurenine and 2-aminobenzoate; the latter is metabolized as known from Azoarcus evansii. Histidine degradation is analogous to the Pseudomonas-type Hut pathway via N-formyl-l-glutamate. For threonine, only one of the three genome-predicted degradation pathways (employing threonine 3-dehydrogenase) is used. Proteins of the individual peripheral degradation sequences in Ph.inhibensDSM 17395 were apparently substrate-specifically formed contrasting the non-modulated TCA cycle enzymes. Comparison of genes for the reconstructed amino acid degradation network in Ph.inhibensDSM 17395 across 27 other complete genomes of Roseobacter clade members revealed most of them to be widespread among roseobacters

The marine bacterium Phaeobacter inhibens secures external ammonium by rapid buildup of intracellular nitrogen stocks.

Reduced nitrogen species are key nutrients for biological productivity in the oceans. Ammonium is often present in low and growth-limiting concentrations, albeit peaks occur during collapse of algal blooms or via input from deep sea upwelling and riverine inflow. Autotrophic phytoplankton exploit ammonium peaks by storing nitrogen intracellularly. In contrast, the strategy of heterotrophic bacterioplankton to acquire ammonium is less well understood. This study revealed the marine bacterium Phaeobacter inhibens DSM 17395, a Roseobacter group member, to have already depleted the external ammonium when only ~1/3 of the ultimately attained biomass is formed. This was paralleled by a three-fold increase in cellular nitrogen levels and rapid buildup of various nitrogen-containing intracellular metabolites (and enzymes for their biosynthesis) and biopolymers (DNA, RNA and proteins). Moreover, nitrogen-rich cells secreted potential RTX proteins and the antibiotic tropodithietic acid, perhaps to competitively secure pulses of external ammonium and to protect themselves from predation. This complex response may ensure growing cells and their descendants exclusive provision with internal nitrogen stocks. This nutritional strategy appears prevalent also in other roseobacters from distant geographical provenances and could provide a new perspective on the distribution of reduced nitrogen in marine environments, i.e. temporary accumulation in bacterioplankton cells