An SDS-PAGE reverse-phase metaproteomics approach for polar desert and ornithogenic soils

Soils are the most biodiverse habitat on earth. Soil micro-organisms mediate global biogeochemical cycles and play vital roles as both pathogens and commensal organisms when interacting with plants, animals and humans. Meta-proteomics offers a promising but underutilised approach to uncovering the functional capacity of soil microbial communities. The potential of meta-proteomics is particularly exciting within low-biomass soils, particularly those in cold deserts, as survival mechanisms under the extreme conditions remains poorly understood. Despite advances in mass spectroscopy platforms, balancing sample purity with yield in soils has posed significant challenges. Here, we address these challenges by optimizing a protein extraction protocol through comparisons of Sodium Dodecyl-Sulfate Polyacrylamide Gel Electrophoresis (SDS-PAGE), SDS-PAGE with reverse-phase chromatography, filter-cleaned, and phenol-extracted samples. SDS-PAGE with reverse-phase chromatography protein purification method was applied to soils from Shirley Island (Antarctica) and Station Nord (Greenland), revealing distinct microbial community dynamics. In Shirley Island soils, algae-bacteria interactions were identified as central to carbon and nitrogen cycling. For hydrocarbon contaminated soils from Station Nord, hydrocarbon presence did not hinder protein discovery, resolving key functional differences, particularly those relating to stress responses and metabolic adaptations. Our low-biomass protein purification protocol proved effective across the studied polar environments, demonstrating its utility for future studies in pristine and contaminated soils worldwide. These findings underscore the value of meta-proteomics in advancing our understanding of microbial roles in extreme environments and their geochemical impacts.