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Organic solvent tolerant oxidoreductases are significant for both scientific research and biomanufacturing. However, it is really challenging to obtain oxidoreductases due to the shortages of natural resources and the difficulty to obtained it via protein modification. This review summarizes the recent advances in gene mining and structure-functional study of oxidoreductases from extremophiles for non-aqueous reaction systems. First, new strategies combining genome mining with bioinformatics provide new insights to the discovery and identification of novel extreme oxidoreductases. Second, analysis from the perspectives of amino acid interaction networks explain the organic solvent tolerant mechanism, which regulate the discrete structure-functional properties of extreme oxidoreductases. Third, further study by conservation and co-evolution analysis of extreme oxidoreductases provides new perspectives and strategies for designing robust enzymes for an organic media reaction system. Furthermore, the challenges and opportunities in designing biocatalysis non-aqueous systems are highlighted.