Evidence for participation of transglutaminase in receptor-mediated endocytosis.
We report evidence that the enzyme transglutaminase (glutaminyl-peptide gamma-glutamyltransferase; R-glutaminyl-peptide:amine gamma-glutamyltransferase, EC2.3.2.13) participates in receptor-mediated endocytosis. Clustering and internalization of rhodamine-labeled alpha 2-macroglobulin (R alpha 2 M) in normal rat kidney (NRK) cells is inhibited by a wide spectrum of compounds that inhibit transglutaminases, including that from NRK cells. The pattern of clustering inhibition resembles the pattern of transglutaminase inhibition as follows: (i) The most potent transglutaminase inhibitors are dansylcadaverine and the transglutaminase-directed affinity label N-benzyloxy-carbonyl-5-diazo-4-oxonorvaline p-nitrophenyl ester; these were also the most potent inhibitors of clustering and internalization of R alpha 2M. (ii) The inhibition of clustering of R alpha 2M occurs in the same concentration range as that required for transglutaminase inhibition. (iii) Linear primary amines are more effective blockers than the iso-chain primary amines. (iv) The transglutaminase affinity label N-benzyloxycarbonyl-5-diazo-4-oxonorvaline p-nitrophenyl ester irreversibly inhibits a significant fraction of the NRK transglutaminase and the clustering and internalization of R alpha 2M. A closely related compound, N-trifluoroacetyl-6-diazo-5-oxonorleucine ethyl ester, does not significantly inhibit transglutaminase or clustering and internalization. (v) Clustering and internalization is inhibited 10-fold more effectively by the heptapeptide Ac-Gly2-LLeu-LLys-Gly3 than by the heptapeptides Ac-Gly2-LLeu-DLys-Gly3 or AcGly3-DLys-DLeu-Gly2. This is the pattern of stereospecificity for the inhibition of purified transglutaminases.
D000345:Affinity Labels; D000818:Animals; D002460:Cell Line; D002463:Cell Membrane Permeability; D004121:Dimethyl Sulfoxide / Q000494:pharmacology; D004705:Endocytosis; D007700:Kinetics; D008560:Membrane Fluidity / Q000187:drug effects; D051381:Rats; D011955:Receptors, Drug / Q000378:metabolism*; D012236:Rhodanine; D000511:alpha-Macroglobulins / Q000378:metabolism; D005723:gamma-Glutamyltransferase / Q000037:antagonists & inhibitors / Q000378:metabolism*
D000345:Affinity Labels; D011955:Receptors, Drug; D000511:alpha-Macroglobulins; D012236:Rhodanine; D005723:gamma-Glutamyltransferase; D004121:Dimethyl Sulfoxide