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Structure of Bacterial Extracellular Polymeric Substances at Different pH Values as Determined by SAXS

Iztok Dogsa ^* , Manfred Kriechbaum , David Stopar  and Peter Laggner 

^* Laboratory for Biophysics, Jozef Stefan Institute, Ljubljana, Slovenia; Biotechnical Faculty, Department of Food Technology, Laboratory of Microbiology, University of Ljubljana, Ljubljana, Slovenia; and Institute of Biophysics and X-Ray Structure Research, Graz, Austria

Correspondence: Address reprint requests to Peter Laggner, E-mail: peter.laggner{at}oeaw.ac.at.

Extracellular polymeric substances (EPS) play an important role in cell aggregation, cell adhesion, and biofilm formation, and protect cells from a hostile environment. The EPS was isolated by trichloroacetic acid/ethanol extraction from broth culture of a marine bacterium isolate. The EPS was composed of glucose and galactose as determined by HPLC and TLC; the protein content was on average 15 ± 5% of EPS dry mass. The solution structure of EPS at different values of pH was revealed by small-angle x-ray scattering. Scattering curves of EPS solutions (0.4%, w/v) consistently showed two nearly linear log-log regions with slopes a and b in the q-ranges from 0.06 nm^–1 to 0.26 nm^–1, and from 0.27 nm^–1 to 0.88 nm^–1, respectively. Slope a was sensitive to pH changes whereas slope b was not. The observed sensitivity to pH was not a consequence of ionic strength variation with pH, as checked by salt addition. The pH variation causes major rearrangements of EPS structure mainly at length scales above 24 nm. To get a better understanding of the pH effect on EPS structure, the original model proposed by Geissler was refined into a mathematical model that enabled fitting of the experimental scattering curves in the pH range from 0.7 to 11.0. The model describes EPS structure as a network of randomly coiled polymeric chains with denser domains of polymeric chains. The results obtained from the model indicate that dense domains increase in average size from 19 nm at pH 11.0 to 52 nm at pH 0.7. The average distance between the polysaccharide chains at pH 0.7 was 2.3 nm, which indicates a compact EPS structure. Swelling was found to be at a maximum around pH = 8.8, where the average distance between the chains was 4.8 nm.