Abstract: The present invention relates to trypsin variants with improved enzymatic properties, and particularly relates to a mutated trypsin comprising an amino acid substitution at least at two amino acid positions leading to an increased affinity for the nucleophilic substrate and/or at least at two amino acid positions leading to a reduced hydrolysis activity.

Abstract: The invention relates to a mutated trypsin comprising an amino acid substitution both at position K60 and D189, and at least one more amino acid substitution by histidine at position N143 or position E151. Such trypsin mutant has a preferred cleavage site comprising the amino acids Xaa1-Xaa2-His, wherein Xaa1 is L, Y or F and Xaa2 is R or K. The invention also relates to a man-made polypeptide comprising a target peptide and the above cleavage site as well as to a method of producing C-terminally modified target peptides by using this mutated trypsin.

Abstract: The invention relates to a mutated trypsin comprising an amino acid substitution both at position K60 and D189, and at least one more amino acid substitution by histidine at position N143 or position E151. Such trypsin mutant has a preferred cleavage site comprising the amino acids Xaa1-Xaa2-His, wherein Xaa1 is L, Y or F and Xaa2 is R or K. The invention also relates to a man-made polypeptide comprising a target peptide and the above cleavage site as well as to a method of producing C-terminally modified target peptides by using this mutated trypsin.

Abstract: The method for producing peptides, peptide mimetics and proteins is characterized in that an enzyme is used as a biocatalyst together with a peptide mimetic. The native specificity of the enzyme is modified by chemical or genetic manipulation, the substrate mimetic is a carboxyl component with an ester leaving group whose specificity determinant is adapted to the recognition site of the enzyme. The invention also concerns the trypsin variant D189K,K60E and its use as a C-N ligase in segment condensations.

Abstract: The invention relates to a mutated trypsin comprising an amino acid substitution both at position K60 and D189, and at least one more amino acid substitution by histidine at position N143 or position E151. Such trypsin mutant has a preferred cleavage site comprising the amino acids Xaa1-Xaa2-His, wherein Xaa1 is L, Y or F and Xaa2 is R or K. The invention also relates to a man-made polypeptide comprising a target peptide and the above cleavage site as well as to a method of producing C-terminally modified target peptides by using this mutated trypsin.

Abstract: The present invention relates to a method for the synthesis of peptides, peptide mimetics and/or proteins and/or for the selective N-terminal modification of peptides, peptide mimetics and/or proteins, with the steps of: a) providing an amino component, said amino component having at least one amino acid, b) providing a carboxyl component, said carboxyl component having a leaving group on the carboxyl group, and said carboxyl component being a compound having at least one amino acid or a compound having at least one label or reporter group, c) reacting said amino component and said carboxyl component in a reaction medium which has one or more ionic liquids, in the presence of a protease, peptidase and/or hydrolase, to form a peptide bond between the amino component and the carboxyl component with elimination of the leaving group.

Abstract: The invention concerns a method for the regiospecific modification of peptides and proteins with probes and reporter molecules using peptidases in combination with a non-amino acid-like or non-peptide-like substrate mimetic.

Abstract: The method for producing peptides, peptide mimetics and proteins is characterized in that an enzyme is used as a biocatalyst together with a peptide mimetic. The native specificity of the enzyme is modified by chemical or genetic manipulation, the substrate mimetic is a carboxyl component with an ester leaving group whose specificity determinant is adapted to the recognition site of the enzyme. The invention also concerns the trypsin variant D189K,K60E and its use as a C—N ligase in segment condensations.