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Computer-aided design of a catalyst for Edman degradation utilizing substrate-assisted catalysis

ABSTRACT: Molecular biology has been revolutionized by the miniaturization and parallelization of DNA sequencing assays previously performed on bulk samples. Many of these technologies rely on biomolecular reagents to facilitate detection, synthesis, or labeling of samples. To aid in the construction of analogous experimental approaches for proteins and peptides, we have used computer-aided design to engineer an enzyme capable of catalyzing the cleavage step of the Edman degradation. We exploit the similarity between the sulfur nucleophile on the Edman reagent and the catalytic cysteine in a naturally occurring protease to adopt a substrate-assisted mechanism for achieving controlled, step-wise removal of N-terminal amino acids. The ability to expose amino acids iteratively at the N-terminus of peptides is a central requirement for protein sequencing techniques that utilize processive degradation of the peptide chain. While this can be easily accomplished using the chemical Edman degradation, achieving this activity enzymatically in aqueous solution removes the requirement for harsh acid catalysis, improving compatibility with low adsorption detection surfaces, such as those used in single molecule assays.

SUBMITTER: Borgo B

PROVIDER: S-EPMC4380987 | biostudies-literature | 2014 Dec

REPOSITORIES: biostudies-literature

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Computer-aided design of a catalyst for Edman degradation utilizing substrate-assisted catalysis

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