N-terminal glycine-specific protein conjugation catalyzed by microbial transglutaminase.
Here, we report the N-terminal glycine (Gly) residue of a target protein can be a candidate primary amine for site-specific protein conjugation catalyzed by microbial transglutaminase (MTG) from Streptomyces mobaraensis. Gly5-enhanced green fluorescent protein (EGFP) (EGFP with five additional Gly residues at its N-terminus) was cross-linked with Myc-dihydrofolate reductase (DHFR) (DHFR with the myc epitope sequence at its N-terminus) to yield DHFR-EGFP heterodimers. The reactivities of additional peptidyl linkers were investigated and the results obtained suggested that at least three additional Gly residues at the N-terminus were required to yield the EGFP-DHFR heterodimeric form. Site-directed mutagenesis analysis revealed marked preference of MTG for amino acids adjacent to the N-terminal Gly residue involved in the protein conjugation. In addition, peptide-protein conjugation was demonstrated by MTG-catalyzed N-terminal Gly-specific modification of a target protein with the myc epitope peptide.
Tanaka
Tsutomu
T
Department of Chemistry and Biotechnology, School of Engineering, The University of Tokyo, 7-3-1 Hongo, Bunkyo-ku, Tokyo 113-8656, Japan.
Kamiya
Noriho
N
Nagamune
Teruyuki
T
D001483:Base Sequence; D017931:DNA Primers; D016297:Mutagenesis, Site-Directed; D013302:Streptomyces / Q000201:enzymology*; D011503:Transglutaminases / Q000235:genetics / Q000378:metabolism*
D017931:DNA Primers; D011503:Transglutaminases