In situ detection of active transglutaminases for keratinocyte type (TGase 1) and tissue type (TGase 2) using fluorescence-labeled highly reactive substrate peptides.
Transglutaminase is a calcium-dependent enzyme that posttranslationally modifies proteins by cross-linking between glutamine and lysine residues or attachment of a primary amine to specific polypeptide-bound glutamine residues. Eight isozymes play essential roles in various mammalian biological processes. The authors have recently identified 12–amino acid preferred substrate peptide sequences that are highly reactive and act in an isozyme-specific manner. In this study, a rapid, isozyme-specific, and sensitive detection of active keratinocyte type (TGase 1) and tissue type (TGase 2) was successful using fluorescence-labeled peptides. This procedure involved using whole-body sections of a mouse to extensively analyze the tissue distribution of both enzymes that revealed clearly distinct patterns. Strong active TGase 1 was observed in epithelial tissues such as tongue, developing teeth, forestomach, and skin epidermis. Significantly active TGase 2 was observed in various types of tissues as predicted and at particularly higher levels in the intestinal mucosa, muscle membrane, and whole veins in the liver. This manuscript contains online supplemental material at http://www.jhc.org. Please visit this article online to view these materials.
Itoh
Miho
M
Department of Applied Molecular Biosciences, Graduate School of Bioagricultural Sciences, Nagoya University, Chikusa, Japan.
Kawamoto
Tadafumi
T
Tatsukawa
Hideki
H
Kojima
Soichi
S
Yamanishi
Kiyofumi
K
Hitomi
Kiyotaka
K
D000595:Amino Acid Sequence; D000818:Animals; D057075:Enzyme Assays / Q000379:methods*; D005456:Fluorescent Dyes / Q000378:metabolism*; D005629:Frozen Sections; D019204:GTP-Binding Proteins / Q000378:metabolism*; D007527:Isoenzymes / Q000378:metabolism; D008297:Male; D051379:Mice; D009842:Oligopeptides / Q000737:chemistry / Q000378:metabolism*; D009928:Organ Specificity; D000091345:Protein Glutamine gamma Glutamyltransferase 2; D013379:Substrate Specificity; D013997:Time Factors; D011503:Transglutaminases / Q000378:metabolism*
D005456:Fluorescent Dyes; D007527:Isoenzymes; D009842:Oligopeptides; D000091345:Protein Glutamine gamma Glutamyltransferase 2; D011503:Transglutaminases; C105804:transglutaminase 1; D019204:GTP-Binding Proteins