Uptake of Amino Acids and Their Metabolic Conversion into the Compatible Solute Proline Confers Osmoprotection to Bacillus subtilis

Abstract

bThe data presented here reveal a new facet of the physiological adjustment processes through which Bacillus subtilis can derive osmostress protection. We found that the import of proteogenic (Glu, Gln, Asp, Asn, and Arg) and of nonproteogenic (Orn and Cit) amino acids and their metabolic conversion into proline enhances growth under otherwise osmotically unfavorable conditions. Osmoprotection by amino acids depends on the functioning of the ProJ-ProA-ProH enzymes, but different entry points into this biosynthetic route are used by different amino acids to finally yield the compatible solute proline. Glu, Gln, Asp, and Asn are used to replenish the cellular pool of glutamate, the precursor for proline production, whereas Arg, Orn, and Cit are converted into ␥-glutamic semialdehyde/⌬ 1 -pyrroline-5-carboxylate, an intermediate in proline biosynthesis. The import of Glu, Gln, Asp, Asn, Arg, Orn, and Cit did not lead to a further increase in the size of the proline pool that is already present in osmotically stressed cells. Hence, our data suggest that osmoprotection of B. subtilis by this group of amino acids rests on the savings in biosynthetic building blocks and energy that would otherwise have to be devoted either to the synthesis of the proline precursor glutamate or of proline itself. Since glutamate is the direct biosynthetic precursor for proline, we studied its uptake and found that GltT, an Na ؉ -coupled symporter, is the main uptake system for both glutamate and aspartate in B. subtilis. Collectively, our data show how effectively B. subtilis can exploit environmental resources to derive osmotic-stress protection through physiological means. Bacillus subtilis is a resident of the upper layers of the soil and of the rhizosphere, and it can also efficiently colonize root surfaces (1-3). The blueprint of its genome (4) bears the hallmarks of a bacterium that can exploit many plant-produced compounds for its growth. Accordingly, a considerable portion of the coding capacity of the B. subtilis chromosome (5) is devoted to highaffinity import systems (6) that allow the scavenging of a wide spectrum of nutrients. Reoccurring and persisting high osmolarity in the soil ecosystem (7) is a situation in which B. subtilis can take advantage of the import of plant-produced compounds (8-10) for its physiological adjustment to these unfavorable environmental conditions (7, 11).As in many bacterial species (11-13), cellular adaptation of B. subtilis to both sudden and sustained increases in the external osmolarity involves a two-stage process (7,14). It initially encompasses the uptake of large quantities of potassium as an emergency stress reaction to curb water efflux (14, 15) and, subsequently, the replacement of part of this ion by organic osmolytes, such as proline (Pro) and glycine betaine (GB), to decrease the ionic strength of the cytoplasm and to optimize its solvent properties (14,(16)(17)(18)(19). These organic osmolytes, commonly referred to as compatible solutes, are highly compliant with cellular physiolog...

Aspartate deficiency limits peptidoglycan synthesis and sensitizes cells to antibiotics targeting cell wall synthesis in Bacillus subtilis

“…By measuring amino acid content in DSM and LB medium, we propose that the growth defect of the aspB null mutant is due to Asp limitation exacerbated by competition with Glu for Asp uptake. Our results suggest that GltT is the major Glu/Asp importer, and are in agreement with prior work (Zaprasis et al, ), even though our conditions are drastically different (our experiments are done in DSM, a complex medium containing all essential amino acids, whereas Zaprasis et al used a glucose ammonium based minimal medium supplemented with 20 μM of Glu or Asp). Growth of the aspB mutant can be strongly inhibited by Glu, even in the presence of DAP, suggesting that all major Asp importers are sensitive to Glu‐inhibition.…”

“…Glutamate is synthesized by glutamate synthase (also known as glutamine oxoglutarate aminotransferase, GOGAT), encoded by gltAB in B. subtilis , which converts one molecule each of Gln and α‐ketoglutarate into two molecules of Glu. Glutamate can be imported into the cell by GltT (Zaprasis et al, ), GltP (Tolner et al, ) and YveA (Lorca et al, ).…”

Short-Term Exposure of Paddy Soil Microbial Communities to Salt Stress Triggers Different Transcriptional Responses of Key Taxonomic Groups

“…Therefore, our research was fully focused on 2-day exposure to salt stress. In pure-culture transcriptome studies, this exposure time is considered long-term incubation given that microorganisms that are able to cope with moderate and high salt stress exhibit an adaptive transcriptome response within 2 days (Zaprasis et al, 2015; Nagler et al, 2016). …”

“…Depending on the ability of the particular microorganism to cope with salt stress, this early period is followed by an adaptive transcriptome response that is governed by up-regulation of gene expression. This adaptive response occurs within a few hours or days (Zaprasis et al, 2015; Nagler et al, 2016). The key taxonomic groups involved in the methanogenic breakdown of plant polymers significantly differed in their ability to cope with salt stress.…”

Identification of the first glyphosate transporter by genomic adaptation

“…3A). The gltT and gltP genes code for the high-affinity sodium-coupled glutamate/aspartate symporter GltT and for the low-affinity proton/glutamate symporter GltP respectively (Tolner et al, 1995;Zaprasis et al, 2015;Zhao et al, 2018). 2B).…”