Directed co-evolution of interacting protein-peptide pairs by compartmentalized two-hybrid replication (C2HR).

ABSTRACT: Directed evolution methodologies benefit from read-outs quantitatively linking genotype to phenotype. We therefore devised a method that couples protein-peptide interactions to the dynamic read-out provided by an engineered DNA polymerase. Fusion of a processivity clamp protein to a thermostable nucleic acid polymerase enables polymerase activity and DNA amplification in otherwise prohibitive high-salt buffers. Here, we recapitulate this phenotype by indirectly coupling the Sso7d processivity clamp to Taq DNA polymerase via respective fusion to a high affinity and thermostable interacting protein-peptide pair. Escherichia coli cells co-expressing protein-peptide pairs can directly be used in polymerase chain reactions to determine relative interaction strengths by the measurement of amplic

SUBMITTER: Siau JW 

PROVIDER: S-EPMC7736784 | biostudies-literature | 2020 Dec

REPOSITORIES: biostudies-literature