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Updated: Jun 27, 2025

Defining Substrate Specificities for Lipase and Phospholipase Candidates
Published on: November 23, 2016
Machine learning model of the catalytic efficiency and substrate specificity of acyl-ACP thioesterase variants
Fuyuan Jing1,2,3, Keting Chen1,4, Marna D Yandeau-Nelson2,3,4
1Roy J. Carver Department of Biochemistry, Biophysics, and Molecular Biology, Iowa State University, Ames, IA, United States.
Abstract:
Modulating the catalytic activity of acyl-ACP thioesterase (TE) is an important biotechnological target for effectively increasing flux and diversifying products of the fatty acid biosynthesis pathway. In this study, a directed evolution approach was developed to improve the fatty acid titer and fatty acid diversity produced by E. coli strains expressing variant acyl-ACP TEs. A single round of in vitro directed evolution, coupled with a high-throughput colorimetric screen, identified 26 novel acyl-ACP TE variants that convey up to a 10-fold increase in fatty acid titer, and generate altered fatty acid profiles when expressed in a bacterial host strain. These in vitro-generated variant acyl-ACP TEs, in combination with 31 previously characterized natural variants isolated from diverse phylogenetic origins, were analyzed with a random forest classifier machine learning tool. The resulting quantitative model identified 22 amino acid residues, which define important structural features that determine the catalytic efficiency and substrate specificity of acyl-ACP TE.
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