Directed evolution of enzymatic silicon-carbon bond cleavage in siloxanes

Creators: Sarai, Nicholas S.; Fulton, Tyler J.; O'Meara, Ryen L.; Johnston, Kadina E.; Brinkmann-Chen, Sabine; Maar, Ryan R.; Tecklenburg, Ron E.; Roberts, John M.; Reddel, Jordan C. T.; Katsoulis, Dimitris E.; Arnold, Frances H.¹

1. California Institute of Technology

Abstract

Volatile methylsiloxanes (VMS) are man-made, nonbiodegradable chemicals produced at a megaton-per-year scale, which leads to concern over their potential for environmental persistence, long-range transport, and bioaccumulation. We used directed evolution to engineer a variant of bacterial cytochrome P450 BM3 to break silicon-carbon bonds in linear and cyclic VMS. To accomplish silicon-carbon bond cleavage, the enzyme catalyzes two tandem oxidations of a siloxane methyl group, which is followed by putative [1,2]-Brook rearrangement and hydrolysis. Discovery of this so-called siloxane oxidase opens possibilities for the eventual biodegradation of VMS.

Copyright and License

© 2024 The Authors, some rights reserved; exclusive licensee American Association for the Advancement of Science. No claim to original U.S. Government Works. This is an article distributed under the terms of the Science Journals Default License.

Acknowledgement

We thank S. Athavale, S. Bähr, L. F. Caldera, and B. J. Levin for productive discussions and experimental assistance and S. Gao and J. C. Reisenbauer for critical review of the manuscript and data. We thank P. J. Almhjell, A. Das, Y. Long, K. Plotzke, G. Kozerski, D. Eldred, E. Joffre, and D. McNett for productive discussions. We thank N. W. Goldberg for the initial design of custom reaction apparatus. We thank D. VanderVelde for NMR assistance and maintenance of the Caltech Liquids NMR facility and M. Shahgoli for insightful discussions, mass spectrometry assistance, and maintenance of the Caltech CCE Multiuser Mass Spectrometry Laboratory. We thank the Dow Next Generation Instrument Grant. We thank E. Hicks and B. Stoltz for access to distillation equipment.

Funding

This work was supported by the Dow University Partnership Initiative (grants 227027AO and 227027AU). N.S.S. and R.L.O. are each supported by grants from the National Science Foundation Graduate Research Fellowship Program (NSF grant DGE-1745301).

Contributions

Conceptualization: N.S.S., T.J.F., R.L.O., K.E.J., S.B.-C., R.R.M., J.M.R., J.C.T.R., D.E.K., and F.H.A. Methodology: N.S.S., T.J.F., R.L.O., K.E.J., S.B.-C., and R.E.T. Investigation: N.S.S., T.J.F., R.L.O., K.E.J., S.B.-C., R.E.T., and J.M.R. Visualization: N.S.S., T.J.F., and R.L.O. Funding acquisition: F.H.A. Project administration: S.B.-C., R.R.M., J.M.R., J.C.T.R., D.E.K., and F.H.A. Supervision: S.B.-C. and F.H.A. Writing – original draft: N.S.S., T.J.F., R.L.O., D.E.K., and F.H.A. Writing – review and editing: N.S.S., T.J.F., R.L.O., K.E.J., S.B.-C., R.R.M., J.M.R., D.E.K., and F.H.A.

Data Availability

All data are available in the main text or the supplementary materials. Plasmids encoding the enzymes reported in this study are available for research purposes from F.H.A. under a material transfer agreement with the California Institute of Technology.

Conflict of Interest

N.S.S., T.J.F., R.L.O., F.H.A., S.B.-C., R.R.M., J.M.R., D.E.K., and J.C.T.R. are inventors on a patent application describing enzymatic siloxane degradation filed by the Dow Chemical Company and the California Institute of Technology. The authors declare no other competing interests.

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Resource type: Journal Article
Publisher: American Association for the Advancement of Science
Published in: Science, 383(6681), 438-443, ISSN: 0036-8075.
Languages: English