Does a simple micellar system mimic the active site of the computationally-designed retro-aldolase enzyme?


Students: Michael Epperson, Joshua Schmidt


Natural enzymes catalyze chemical reactions with tremendous rate accelerations and exquisite specificities. A long-term goal in enzymology and in protein engineering has been to rationally design new enzymes for chemical reactions. In particular, a recently computationally designed enzyme was found to catalyze a retro-aldol reaction with rate acceleration, compared to the uncatalyzed reaction in solution, of about five orders of magnitude [1].


Biochemical characterization of the computationally designed retroaldolase showed that functional groups predicted by computation to be important for catalysis were in fact contributing only 30-fold to the rate acceleration, and suggested the possibility that most of the catalytic power of the enzyme was instead due to positioning of the substrate within the enzyme’s active site [2]. Indeed, the use of binding energies to fix substrates in a reactive conformation has been shown to be an important catalytic strategy used by enzymes.


Because the substrate of the retro-aldol enzyme is a naphthyl derivative, 4-hydroxy-4-(6-methoxy-2-naphthyl)-2-butanone (HMNB), it is possible that sequestering the apolar naphthyl ring from aqueous solution and positioning it in an environment that contains a nucleophilic lysine residue, might account for most of the 105-fold rate acceleration of the computationally designed retroaldolase. In this work we will test this hypothesis by measuring the rate of the amine-catalyzed retro-aldol reaction of HMNB in micelles, amphiphilic compounds that can self-assemble so that a relatively hydrophobic core is formed even in polar water. Positioning HMNB in this core might result in significant rate acceleration.

[1] Jiang, L.; Althoff, E. A.; Clemente, F. R.; Doyle, L.; Rothlisberger, D.; Zanghellini, A.; Gallaher, J. L.; Betker, J. L.; Tanaka, F.; Barbas, C. F., 3rd; Hilvert, D.; Houk, K. N.; Stoddard, B. L.; Baker, D. Science 2008, 319, 1387.


[2] Lassila, J. K.; Baker, D.; Herschlag, D. Proc Natl Acad Sci U S A 2010, 107, 4937.