TY - JOUR
T1 - Bildung von oligomeren Formen des Insulinrezeptors durch intramolekularen Disulfidaustausch. Beteiligung von maskierten SH-Gruppen
T2 - Involvement of Masked Sulfhydryl Groups
AU - Krämer, H.
AU - Deger, A.
AU - Koch, R.
AU - Rapp, R.
AU - Hinz, M.
AU - Weber, U.
N1 - Funding Information:
This work was supported by the Deutsche gemeinschaft Grant We 524/8.
PY - 1987
Y1 - 1987
N2 - Insulin receptors from rat liver membranes were labelled with a 125I-labelled photo-reactive insulin analogue or by iodination using lactoperoxidase and analysed by sodium dodecyl sulfate Polyacrylamide gel electrophoresis. Under nonreducing conditions different receptor forms with Mr 400 000 (α2β2), 360000 (α2/ββ) 330 000 (α2 β β), 320 000 (α2β), 280 000 (α2 β') 240000 (α2), 210 000 (αß), 165 000 (αß') and 115 000 (ce) were detected. The subunit composition of these receptor forms was determined by two-dimensional sodium dodecyl sulfate Polyacrylamide gel electrophoresis in the absence and presence of dithioerythritol. During denaturation in sodium dodecyl sulfate in the absence of reductants, the Mr 400 000 receptor form (α2β2) was converted into the Mr 320 000 (α2 β) and Mr 240 000 (α2) receptor form. This conversion was prevented either by N-ethylmaleimide, oxidants, or low pH. In contrast, alkylation of the receptor with N-ethylmaleimide under non-denaturing conditions did not prevent the appearance of intermediate-sized receptor forms. Furthermore, the inhibition of receptor cleavage by N-ethylmaleimide during denaturation was also observed when the amount of free sulfhydryl groups was reconstituted by the addition of an unlabelled and non-alkylated receptor sample to the alkylated and photoaffinity-la-belled receptor. These results suggest, that the generation of different oligomeric receptor forms detected by sodium dodecyl sulfate Polyacrylamide gel electrophoresis is due at least in part to the cleavage of one or both β-subunits from the insulin receptor. This cleavage is the result of an intramolecular sulfhydryl-disulfide exchange involving masked sulfhydryl groups of the receptor itself which are uncovered by sodium dodecyl sulfate.
AB - Insulin receptors from rat liver membranes were labelled with a 125I-labelled photo-reactive insulin analogue or by iodination using lactoperoxidase and analysed by sodium dodecyl sulfate Polyacrylamide gel electrophoresis. Under nonreducing conditions different receptor forms with Mr 400 000 (α2β2), 360000 (α2/ββ) 330 000 (α2 β β), 320 000 (α2β), 280 000 (α2 β') 240000 (α2), 210 000 (αß), 165 000 (αß') and 115 000 (ce) were detected. The subunit composition of these receptor forms was determined by two-dimensional sodium dodecyl sulfate Polyacrylamide gel electrophoresis in the absence and presence of dithioerythritol. During denaturation in sodium dodecyl sulfate in the absence of reductants, the Mr 400 000 receptor form (α2β2) was converted into the Mr 320 000 (α2 β) and Mr 240 000 (α2) receptor form. This conversion was prevented either by N-ethylmaleimide, oxidants, or low pH. In contrast, alkylation of the receptor with N-ethylmaleimide under non-denaturing conditions did not prevent the appearance of intermediate-sized receptor forms. Furthermore, the inhibition of receptor cleavage by N-ethylmaleimide during denaturation was also observed when the amount of free sulfhydryl groups was reconstituted by the addition of an unlabelled and non-alkylated receptor sample to the alkylated and photoaffinity-la-belled receptor. These results suggest, that the generation of different oligomeric receptor forms detected by sodium dodecyl sulfate Polyacrylamide gel electrophoresis is due at least in part to the cleavage of one or both β-subunits from the insulin receptor. This cleavage is the result of an intramolecular sulfhydryl-disulfide exchange involving masked sulfhydryl groups of the receptor itself which are uncovered by sodium dodecyl sulfate.
KW - Insulin receptor oligomeric forms
KW - effect of insulin
KW - photoaffinity labelling
KW - subunit composition
KW - sulfhydryl-disulfid exchange
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U2 - 10.1515/bchm3.1987.368.1.471
DO - 10.1515/bchm3.1987.368.1.471
M3 - Article
C2 - 3304334
AN - SCOPUS:0023340620
SN - 0177-3593
VL - 368
SP - 471
EP - 480
JO - Biological Chemistry Hoppe-Seyler
JF - Biological Chemistry Hoppe-Seyler
IS - 1
ER -