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

Protein Science. 2014; 
Benjamin Borgo James J. Havrane
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Custom Vector Construction … A synthetic gene encoding the engineered enzyme was purchased from GenScript (Piscataway, NJ) and cloned into a pET42(a) (Novagen) expression vector between the NdeI and XhoI sites, then transformed into E coli, BL‐21(DE3) chemically competent cells … Get A Quote

摘要

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‐termi... More

关键词

protein engineering substrate‐assisted catalysis Edman degradation peptide analysis