Revealing the Compact Structure of Lactic Acid Bacterial Heteroexopolysaccharides by SAXS and DLS

Abstract

Molecular structures of exopolysaccharides are required to understand their functions and the relationships between the structure and physical and rheological properties. Small-angle X-ray scattering and dynamic light scattering were used in conjunction with molecular modeling to characterize solution structures of three lactic acid bacterial heteroexopolysaccharides (HePS-1, HePS-2, and HePS-3). Values of radius of gyration R, cross-sectional radius of gyration R, approximate length L, and hydrodynamic diameter were not directly proportional to the molar mass and indicated the HePSs adopted a compact coil-like rather than an extended conformation. Constrained molecular modeling of 15000 randomized HePS-1 conformers resulted in five best-fit structures with R factor of 3.9-4.6% revealing random coil-like structure. Φ and Ψ angle analysis of glycosidic linkages in HePS-1 structures suggests Galf residues significantly influence the conformation. Ab initio scattering modeling of HePS-2 and HePS-3 gave excellent curve fittings with χ of 0.43 and 0.34 for best-fit models, respectively, compatible with coil-like conformation. The findings disclose solution behavior of HePS relevant for their interactions with biomacromolecules, for example, milk proteins.

Interaction between structurally different heteroexopolysaccharides and β-lactoglobulin studied by solution scattering and analytical ultracentrifugation

“…lactis CNRZ 371 [33], Streptococcus thermophilus EU20 [34] and Streptococcus thermophilus RD534 [35], respectively and measured their interactions with βlactoglobulin (BLG) using small-angle X-ray scattering (SAXS), dynamic light scattering (DLS) and analytical ultracentrifugation (AUC). These methods are well developed and have previously been used to examine solution structures of HePSs [36], structural evolution of metastable protein aggregates [37], heat-and salt-induced aggregation of BLG [38,39], as well as zinc-and polyanion-induced selfassociation of complement factor H [40,41]. The present data provide novel information on associative interactions between a collection of structure-determined HePSs and BLG.…”

“…backbone, which make HePS-2 more flexible to adopt a compact conformation in solution [36]. These data suggest that a high degree of branching may promote optimal binding with proteins.…”

“…HePS-1, HePS-2 and HePS-5 were produced in 10% skimmed milk medium by Lactobacillus delbrueckii ssp. bulgaricus NCIMB 702483 (NCIMB, Aberdeen, Scotland, United Kingdom), Lactobacillus rhamnosus GG (ATCC 53103) (ATCC, Manassas, VA, USA) and Streptococcus thermophilus EU20 (kind gift of Prof. Luc De Vuyst, Vrije Universiteit Brussels, Belgium), respectively, and were purified from the culture medium as we have described previously [36].…”

“…Molecular weights of HePS-1HePS-6 were determined by size-exclusion chromatography (SEC) as described [36], using a solvent delivery system (LC-10AD), autosampler (SIL-10A), RI detector (RID-10A; all Shimadzu, Kyoto, Japan) and a size-exclusion column (Shodex OH-PK SB-805 HQ 300 × 8 mm, Showa Denko, Tokyo, Japan). Dextran standards of M w 4.…”

“…For instance, the associative interactions between BLG and HePS-1 can be attributed to the presence of α-1→2 and α-1→3 linkages in the repeating units and the degree of branching involving α-1→3, β-1→3 and β-1→4 linkages, conferring semi-flexible behavior to HePS-1, allowing to adopt a local helical-like structure with a cluster of branches on one side of the backbone, with a similar cluster forming on the other side of the backbone. Thus HePS-1 structure has a compact conformation [36] that is preformed for optimal binding with proteins. Similarly, the results of the present study also showed that BLG aggregated strongly with the HePS containing a high degree of…”