7RNQ

Holo structure of engineered TrpB, 2B9-H275E, from Pyrococcus furiosus in the extended-open conformation

Summary for 7RNQ

• Entry DOI: 10.2210/pdb7rnq/pdb
• Descriptor: Tryptophan synthase beta chain 1, PHOSPHATE ION, SODIUM ION, ... (4 entities in total)
• Functional Keywords: plp, tryptophan synthase, directed evolution, biosynthetic protein
• Biological source: Pyrococcus furiosus
• Total number of polymer chains: 4
• Total formula weight: 174910.60

Authors

Higgins, P.M.,Buller, A.R. (deposition date: 2021-07-29, release date: 2022-08-03, Last modification date: 2023-11-15)

Primary citation

McDonald, A.D.,Higgins, P.M.,Buller, A.R.
Substrate multiplexed protein engineering facilitates promiscuous biocatalytic synthesis.
Nat Commun, 13:5242-5242, 2022

PubMed Abstract: Enzymes with high activity are readily produced through protein engineering, but intentionally and efficiently engineering enzymes for an expanded substrate scope is a contemporary challenge. One approach to address this challenge is Substrate Multiplexed Screening (SUMS), where enzyme activity is measured on competing substrates. SUMS has long been used to rigorously quantitate native enzyme specificity, primarily for in vivo settings. SUMS has more recently found sporadic use as a protein engineering approach but has not been widely adopted by the field, despite its potential utility. Here, we develop principles of how to design and interpret SUMS assays to guide protein engineering. This rich information enables improving activity with multiple substrates simultaneously, identifies enzyme variants with altered scope, and indicates potential mutational hot-spots as sites for further engineering. These advances leverage common laboratory equipment and represent a highly accessible and customizable method for enzyme engineering.

PubMed: 36068220
DOI: 10.1038/s41467-022-32789-w

Experimental method

X-RAY DIFFRACTION (2.102 Å)