Journal
FEBS LETTERS
Volume 579, Issue 10, Pages 2092-2096Publisher
WILEY
DOI: 10.1016/j.febslet.2005.02.064
Keywords
site-specific protein conjugation; N-terminal-specific protein modification; enzymatic cross-linking reaction; posttranslational modification; transglutaminase
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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. (c) 2005 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.
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