Related Experiment Video
Updated: Jul 19, 2026

The Use of a β-lactamase-based Conductimetric Biosensor Assay to Detect Biomolecular Interactions
Published on: February 1, 2018
Structure-guided SCHEMA recombination of distantly related beta-lactamases
Michelle M Meyer1, Lisa Hochrein, Frances H Arnold
1Biochemistry and Molecular Biophysics, California Institute of Technology Mail Code 210-21, California Institute of Technology Mail Code 210-41, Pasadena, CA 91125, USA.
Abstract:
We constructed a library of beta-lactamases by recombining three naturally occurring homologs (TEM-1, PSE-4, SED-1) that share 34-42% sequence identity. Most chimeras created by recombining such distantly related proteins are unfolded due to unfavorable side-chain interactions that destabilize the folded structure. To enhance the fraction of properly folded chimeras, we designed the library using SCHEMA, a structure-guided approach to choosing the least disruptive crossover locations. Recombination at seven selected crossover positions generated 6561 chimeric sequences that differ from their closest parent at an average of 66 positions. Of 553 unique characterized chimeras, 111 (20%) retained beta-lactamase activity; the library contains hundreds more novel beta-lactamases. The functional chimeras share as little as 70% sequence identity with any known sequence and are characterized by low SCHEMA disruption (E) compared to the average nonfunctional chimera. Furthermore, many nonfunctional chimeras with low E are readily rescued by low error-rate random mutagenesis or by the introduction of a known stabilizing mutation (TEM-1 M182T). These results show that structure-guided recombination effectively generates a family of diverse, folded proteins even when the parents exhibit only 34% sequence identity. Furthermore, the fraction of sequences that encode folded and functional proteins can be enhanced by utilizing previously stabilized parental sequences.
Related Concept Videos
Conservative Site-specific Recombination and Phase Variation
The recognition sites for Cre recombinase called LoxP...
Inhibitors of Gram-positive Cell Wall Synthesis
Modern Molecular Taxonomy
Ligand Binding and Linkage
Allosteric Proteins-ATCase
Aspartate transcarbamoylase (ATCase) is a cytosolic enzyme that catalyzes the condensation of L-aspartate and carbamoyl phosphate to N-carbamoyl-L-aspartate. This reaction is the first step in pyrimidine biosynthesis. UTP and CTP, the end products of the pyrimidine synthesis pathway,...
Clinical Significance of Antibiotic Resistance

