Esterase activity of bovine serum albumin up to 160 °C: A new benchmark for biocatalysis

Albumins are highly conserved proteins that carry out a multitude of physiological functions and exhibit a broad range of catalytic activities. Moreover, amino acid sequence comparisons of the albumin multigene family indicate that albumins diverged from a common ancestor. Here we report that bovine serum albumin (BSA) can catalyze ester hydrolysis at high temperatures (as high as 160 °C) well beyond the temperature limits reported for enzymatic catalysis, including for enzymes from known hyperthermophiles. Furthermore, BSA exhibited a ∼133-fold increase in its turnover number (kcat) toward p-nitrophenyl palmitate from 70 to 150 °C. When BSA was incubated for 1 h at 150 °C in the presence of 25 mM SDS, it retained complete esterase activity, indicating that a catalytically competent orientation of amino acid residues exists in the denatured or partially unfolded protein. However, esterase activity diminished to ∼50% upon disruption of the protein's disulfide bridges and disappeared completely when BSA was digested by proteases. These results point to a new standard of robustness for biocatalytic activity at high temperatures. Catalytic activity and promiscuity at very high temperatures could have been advantageous to enzymes in primitive organisms evolving in hot environments, making BSA an intriguing model for early enzymes.