Total Chemical Synthesis of the Enzyme Sortase A_ΔN59 with Full Catalytic Activity

Abstract

Abstract:The enzyme sortase A is a ligase which catalyzes transpeptidation reactions.

Engineering sortase A by screening a second‐generation library using phage display

“…Several research approaches have focused on engineering sortase A in order to understand its reaction mechanism and to extend the applicability of SML Directed evolution has been used for altering the substrate selectivity of sortase and for enhancing the catalytic properties of the enzyme We have previously reported a phage display system for evolving the selectivity of leucine residue within the LPxTG sorting motif . A library of sortase mutants with randomized β 6/ β 7 loop, which is predicted to interact with the N‐terminal residue of the sorting motif, was generated and screened for enzymes recognizing an FPxTG motif by phage display.…”

Postligation-Desulfurization: A General Approach for Chemical Protein Synthesis

“…Using this highly versatile free-radical cysteine reduction protocol, a serious of proteins with complex structures and up to 306 amino acid residues have been synthesized, including human glycosyl-interferon-β [55], orf virus entry fusion proteins ORFV036 and 049 [56], enzyme sortase A ΔN59 [57], antifreeze proteins (AFPs) and antifreeze glycoproteins (AFGPs) [58,59], ribosomal protein S25 (RpS25) [60], α-synuclein (α-syn) [61,62], granulocyte-macrophage colonystimulating factor (GM-CSF) [57], etc. Among these protein syntheses, one of the most fantastic examples is the total synthesis of homogeneous erythropoietin (EPO).…”

Sortagging: A Robust and Efficient Chemoenzymatic Ligation Strategy

“…[125] Recently, Kent et al reported the total chemical synthesis of an enzymatically active sortase A that enables a more extensive engineering of the enzyme. [126] Due to its specificity the sortase is also applicable to orthogonal labeling strategies. Apart from all these advantages over other conventional labeling approaches, several drawbacks including the reversibility of the transpeptidation have to be considered.…”