High-molecular-weight human epidermal transglutaminase.
Human stratum corneum was extracted in Tris-HCl containing EDTA and phenylmethylsulfonyl fluoride, separated on sodium dodecyl sulfate-polyacrylamide gel electrophoresis, transblotted to nitrocellulose papers and reacted with rabbit antihuman epidermal transglutaminase (ETG) antibody. Protein-bound antibody was detected with a multistep peroxidase procedure. Proteins with a molecular weight of 50,000 (50kDa) and 72,000 daltons (72kDa) were stained when anti-ETG was used and not when second antibody alone or sera from nonimmunized animals were used. When ETG was treated with trypsin or organic solvents, there was no alteration in the mobility of the 50kDa ETG band, but there was complete disappearance of the 72kDa band. Antibody that bound 72kDa protein, when eluted from the blot, reacted with both 50kDa and 72kDa proteins; similarly, antibody that bound to the 50kDa protein, when eluted from the blot, reacted with both the 50kDa and 72kDa proteins. Partially purified 72kDa ETG activity was increased (3 to 16 times control levels) after heating at 56 degrees C in the presence of calcium and dithiothreitol or by treatment with trypsin. These studies, in conjunction with the previous studies of ETG activation, are consistent with there being two forms of ETG. The different forms may play a role in regulating enzyme activity.
Negi
M
M
Colbert
M C
MC
Goldsmith
L A
LA
D000217:Acyltransferases / Q000032:analysis* / Q000276:immunology / Q000302:isolation & purification; D000818:Animals; D000906:Antibodies / Q000276:immunology; D002851:Chromatography, High Pressure Liquid; D003429:Cross Reactions; D004591:Electrophoresis, Polyacrylamide Gel; D004789:Enzyme Activation; D006801:Humans; D007124:Immunoenzyme Techniques; D008970:Molecular Weight; D011817:Rabbits; D012867:Skin / Q000201:enzymology*; D011503:Transglutaminases
D000906:Antibodies; D000217:Acyltransferases; D011503:Transglutaminases