Ligase fusion proteins and application thereof

An immobilized ligase, comprising a ligase fusion protein immobilized to a support, wherein the ligase fusion protein consists of a ligase, a mutant haloalkane dehalogenase or a variant thereof comprising the amino acid sequence of SEQ ID NO:

• 28 or an amino acid sequence having a sequence identity of at least 95% thereto, and optionally a linker peptide, wherein the ligase, the mutant haloalkane dehalogenase or the variant thereof, and optionally the linker peptide are fused by a covalent bond, wherein the ligase has an isoelectric point (pI) of about 7.5 to about 10.0,
• the mutant haloalkane dehalogenase or the variant thereof has an isoelectric point of about 4.5 to about 5.0, and
• the pI of the ligase fusion protein is about 2.0 to about 4.5 pH units lower than that of the ligase, wherein the ligase is a sortase, and wherein the support comprises a haloalkyl linker such that the ligase fusion protein is immobilized on the support through covalent interaction between the haloalkyl linker and the mutant haloalkane dehalogenase or the variant thereof.