CAZyme fold architecture is conserved between disparate environments despite extreme sequence divergence

Microbial carbohydrate-active enzymes (CAZymes) underpin carbon cycling across Earth’s ecosystems; however, how contrasting environments shape CAZyme diversity and structural conservation remains poorly understood. Here, we applied shotgun metagenomics to compare the carbohydrate-degradation potential of two functionally prolific but physicochemically opposed ecosystems: the alkaline-saline soda lakes of the East African Rift Valley and the anaerobic ruminant gut. From 34 metagenomes (12 soda lake and 22 rumen), we recovered 371 quality-filtered metagenome-assembled genomes, of which 84% of soda lake and 52% of rumen MAGs represented novel species. Rumen communities, dominated by Bacteroidota, Fibrobacterota, and Bacillota, exhibited significantly higher taxonomic diversity and were enriched in carbohydrate catabolism and fermentation pathways. Soda lake communities, dominated by Pseudomonadota, displayed greater evolutionary divergence (lower RED scores) and were enriched in pH homeostasis, oxidative and osmotic stress, sulfur cycling, and carbon fixation pathways. To assess whether structural conservation persists despite extreme sequence divergence, we predicted three-dimensional structures for 12 representative enzymes from six glycoside hydrolase families (GH1, GH3, GH5_11, GH9, GH10, and GH28) using AlphaFold 3. All 12 structures adopted canonical GH family folds with high confidence (pTM 0.75–0.97). These results demonstrate that environmental selection drives distinct taxonomic and functional strategies for carbon processing while preserving three-dimensional CAZyme architecture, positioning soda lake and rumen metagenomes as complementary reservoirs for bioprospecting industrially relevant enzymes.