The glutamine residues reactive in transglutaminase-catalyzed cross-linking of involucrin.
The protein involucrin, synthesized by human keratinocytes, contains 585 amino acids, largely in the form of 10 amino acid repeats, each containing glutamines in 3 conserved positions. Involucrin is a substrate for the keratinocyte transglutaminase and is labeled by the cosubstrate amine, glycine ethyl ester. Study of tryptic peptides of involucrin shows that a single glutamine (residue 496), located 89 residues from the C-terminal end, is preferentially labeled by the enzyme. Additional glutamine residues become reactive when the molecule is fragmented. The C-terminal end, isolated as a cyanogen bromide fragment of 275 residues, is labeled equally at 2 glutamine residues. The polypeptide containing residues 148 to 280 accepts practically no amine while in intact involucrin but as a free fragment is labeled at multiple glutamine residues. It is concluded that the C-terminal and N-terminal ends of the protein are directive influences in that they suppress the reactivity of a number of glutamine residues in the intact molecule, leaving one glutamine highly preferred by the transglutaminase.
Simon
M
M
Department of Cellular and Molecular Physiology, Harvard Medical School, Boston, Massachusetts 02115.
Green
H
H
D000595:Amino Acid Sequence; D002478:Cells, Cultured; D003432:Cross-Linking Reagents; D003488:Cyanogen Bromide; D003600:Cytosol / Q000378:metabolism; D004817:Epidermis / Q000378:metabolism; D005973:Glutamine; D006801:Humans; D007633:Keratins / Q000378:metabolism; D010446:Peptide Fragments / Q000032:analysis; D011498:Protein Precursors / Q000378:metabolism*; D011503:Transglutaminases / Q000378:metabolism*
D003432:Cross-Linking Reagents; D010446:Peptide Fragments; D011498:Protein Precursors; D005973:Glutamine; C032253:involucrin; D007633:Keratins; D011503:Transglutaminases; D003488:Cyanogen Bromide