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.

Provided is a polypeptide combination, a targeting component thereof comprising a shielding peptide, a cleavable part, an antigen-binding part and a first intein fragment, the shielding peptide and the antigen-binding part being connected by means of the cleavable part, and the antigen-binding part being directly or indirectly connected to the first intein fragment; a toxin component thereof comprises a second intein fragment and a toxin, and the second intein fragment is directly or indirectly connected to the toxin; the targeting component and the toxin component form an immunoconjugate by means of the interactive action between the first intein fragment and the second intein fragment; in the immunoconjugate, the shielding peptide and the antigen-binding part are connected by means of the cleavable part, and the antigen-binding part is connected to the toxin. Also provided are a preparation method and a pharmaceutical use for the polypeptide combination.

The present invention relates to a signal sequence for expressing a CRM197 protein in Escherichia coli and secreting same into the periplasm, and a use thereof, and more specifically, to: a signal sequence for expressing a CRM197 protein; a nucleic acid for coding the signal sequence; a nucleic acid construct or expression vector comprising the nucleic acid and a CRM197 protein gene; a recombinant microorganism having the nucleic acid construct or expression vector introduced therein; and a CRM197 protein production method comprising a step for culturing the recombinant microorganism. According to the present invention, a CRM197 protein having the same physicochemical/immunologic properties as the protein isolated from the parent bacteria may be expressed even in regular Escherichia coli of which a redox potential is not adjusted, and a CRM197 protein having high periplasmic secretion efficiency may be produced even without shifting the pH of a culture medium in order to increase secretion into the periplasm, and thus the present invention is very useful in CRM197 protein production.

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.

The present disclosure relates to a microorganism producing L-lysine, and a method for producing L-lysine using the same.

The present disclosure relates to a microorganism producing L-lysine, and a method for producing L-lysine using the same.

The present invention relates to a method for producing a periplasmic form of SEQ ID NO: 12, to the expression vector encoding SEQ ID NO: 12 and signal sequence SEQ ID NO: 2, enabling the targeting of SEQ ID NO: 12 towards the periplasmic space, as well as to the strain transformed by the expression vector.