Engineered repeat proteins as scaffolds to assemble multi-enzyme systems for efficient cell-free biosynthesis.

ABSTRACT: Multi-enzymatic cascades with enzymes arranged in close-proximity through a protein scaffold can trigger a substrate channeling effect, allowing for efficient cofactor reuse with industrial potential. However, precise nanometric organization of enzymes challenges the design of scaffolds. In this study, we create a nanometrically organized multi-enzymatic system exploiting engineered Tetrapeptide Repeat Affinity Proteins (TRAPs) as scaffolding for biocatalysis. We genetically fuse TRAP domains and program them to selectively and orthogonally recognize peptide-tags fused to enzymes, which upon binding form spatially organized metabolomes. In addition, the scaffold encodes binding sites to selectively and reversibly sequester reaction intermediates like cofactors via electrostatic interaction

SUBMITTER: Ledesma-Fernandez A 

PROVIDER: S-EPMC10160029 | biostudies-literature | 2023 May

REPOSITORIES: biostudies-literature