Abstract
Microbial decomposition of organic matter is an essential process in the global carbon cycle. The soil bacteria Pseudopedobacter saltans and Flavobacterium johnsoniae are both able to degrade complex organic molecules, but it is not fully known how their membrane structures are adapted to their environmental niche. The membrane lipids of these species were extracted and analyzed using high performance liquid chromatography-electrospray ionization/ion trap/mass spectrometry (HPLC-ESI/IT/MS) and high resolution accurate mass/mass spectrometry (HRAM/MS). Abundant unknown intact polar lipids (IPLs) from P. saltans were isolated and further characterized using amino acid analysis and two dimensional nuclear magnetic resonance (NMR) spectroscopy. Ornithine IPLs (OLs) with variable (hydroxy) fatty acid composition were observed in both bacterial species. Lysine-containing IPLs (LLs) were also detected in both species and were characterized here for the first time using HPLC-MS. Novel LLs containing hydroxy fatty acids and novel hydroxylysine lipids with variable (hydroxy) fatty acid composition were identified in P. saltans. The confirmation of OL and LL formation in F. johnsoniae and P. saltans and the presence of OlsF putative homologs in P. saltans suggest the OlsF gene coding protein is possibly involved in OL and LL biosynthesis in both species, however, potential pathways of OL and LL hydroxylation in P. saltans are still undetermined. Triplicate cultures of P. saltans were grown at three temperature/pH combinations: 30°C/pH 7, 15°C/pH 7, and 15°C/pH 9. The fractional abundance of total amino acid containing IPLs containing hydroxylated fatty acids was significantly higher at higher temperature, and the fractional abundance of lysine-containing IPLs was significantly higher at lower temperature and higher pH. These results suggest that these amino acid-containing IPLs, including the novel hydroxylysine lipids, could be involved in temperature and pH stress response of soil bacteria.
Highlights
Intact polar lipids (IPLs) are useful biomarker molecules because their structures can be specific to microbial taxa or environmental conditions (Sturt et al, 2004; Schubotz et al, 2009)
We report the intact polar lipids (IPLs) content of P. saltans, and F. johnsoniae including the structural elucidation of novel amino acid-containing lipids and discuss the potential genes and environmental factors that influence their production in P. saltans
Two low abundance IPLs with Ornithine IPLs (OLs)-like fragmentation were observed at retention times 27.7 and 28.0 min with apparent protonated molecules ([M+H]+) at m/z values 639 and 625, respectively, making up 4.6% of HPLC-ESI/IT/MS chromatogram base peak area, which we will refer to as group I
Summary
Intact polar lipids (IPLs) are useful biomarker molecules because their structures can be specific to microbial taxa or environmental conditions (Sturt et al, 2004; Schubotz et al, 2009). Marine phytoplankton from multiple regions of the ocean produce non-phosphorus containing lipids in response to phosphorus scarcity (Van Mooy et al, 2009), which demonstrates the importance of microbial membrane modification. IPLs containing the amino acid ornithine as the polar head group (ornithine lipids, OLs) are common phosphorus-free membrane IPLs among bacteria (Figure 1A). OL production increases under phosphorus limitation (Weissenmayer et al, 2002; Gao et al, 2004), and in other microbes the fatty acids of OLs are hydroxylated under thermal or acid stress (Taylor et al, 1998; Rojas-Jimenez et al, 2005; Vences-Guzman et al, 2011). It has been suggested that OLs are important for Gram-negative bacterial outer membrane stability due to their zwitterionic nature (Freer et al, 1996) and essential to maintain a constant level of extracytoplasmic cytochromes in Rhodobacter capsulatus (Aygun-Sunar et al, 2006)
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