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AssigneesThe Regents of the University of California, Board of Regents, The University of Texas System
InventorsRobert T. Clubb, Brendan Rayhan Amer, Janine Y. Fu, Scott McConnell, Hung Ton-That, Chungyu Chang
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Covalently cross-linked pilus polymers displayed on the cell surface of Gram-positive bacteria are assembled by class C sortase enzymes. These pilus-specific transpeptidases located on the bacterial membrane catalyze a two-step protein ligation reaction—first, cleaving the LPXTG motif of one pilin protomer to form an acyl-enzyme intermediate, and second, joining the terminal threonine to the nucleophilic lysine residue residing within the pilin motif of another pilin protomer. Informed by the high-resolution crystal structures of corynebacterial pilus-specific sortase (SrtA) and by developing structural variants of the sortase enzyme whose catalytic pocket has been unmasked by activating mutations, we have developed new reagents capable of forming isopeptide bonds in vitro. The reagents disclosed herein can catalyze ligation of isolated SpaA domains in vitro provide a facile and versatile new platform for protein engineering and bio-conjugation that has major implications for biotechnology.
A method of forming an isopeptide bond in vitro between a threonine residue in a first polypeptide and a lysine residue in a second polypeptide, the method comprising:
A method of forming an isopeptide bond in vitro between a threonine residue in a first polypeptide and a lysine residue in a second polypeptide, the method comprising:
A method of forming an isopeptide bond in vitro between a threonine residue in a first polypeptide and a lysine residue in a second polypeptide, the method comprising:
A method of forming an isopeptide bond in vitro between a threonine residue in a first polypeptide and a lysine residue in a second polypeptide, the method comprising:
Dependencies are inferred from the claim text.