Degradation of cadmium metallothionein in vitro by lysosomal proteases.
The effects of various protease inhibitors on the degradation of cadmium metallothionein (Cd-MT) by lysosomal proteases were studied in vitro. Degradation of Cd-MT was observed after incubation with the lysosomal extracts, but not after incubation with the cytosol or heat-treated lysosomal extracts. After incubation of [35S]-Cd-MT or 109Cd-MT with lysosomal extracts, 35S and 109Cd radioactivity in the MT fraction decreased, while the low molecular weight (LM) fraction increased with time (half life; 3 hr). When EDTA was added to this incubation mixture, most of the MT was degraded within 30 min. Cd in the LM fraction, produced after the incubation of Cd-MT with the lysosomal extracts, was moved to the high molecular weight fraction by the addition of cytosol. Both leupeptin and E-64, which reduced cathepsin B (cysteine protease) activity, inhibited the degradation of Cd-MT by the lysosomal extracts. But pepstatin A, a specific inhibitor of cathepsin D, did not inhibit this degradation. E-64 inhibited degradation, as well as inhibiting cathepsin B activity, in accordance with its concentration in the incubation mixture. The incubation of Cd-MT with purified cathepsin B resulted in its degradation which was inhibited by E-64. These results suggest that Cd-MT may be broken down by the cysteine protease in lysosomes and that the released Cd bound low molecular weight fragment(s) was subsequently transferred to the high molecular weight protein in cytosol.
Min
K S
KS
Department of Nutrition, Kobe-Gakuin University, Japan.
Nakatsubo
T
T
Fujita
Y
Y
Onosaka
S
S
Tanaka
K
K
D000818:Animals; D002104:Cadmium / Q000378:metabolism / Q000494:pharmacology*; D019256:Cadmium Chloride; D002403:Cathepsins / Q000037:antagonists & inhibitors; D003600:Cytosol / Q000187:drug effects / Q000378:metabolism; D007279:Injections, Subcutaneous; D008099:Liver / Q000187:drug effects* / Q000378:metabolism; D008247:Lysosomes / Q000187:drug effects* / Q000378:metabolism; D008297:Male; D008668:Metallothionein / Q000378:metabolism*; D011480:Protease Inhibitors / Q000494:pharmacology*; D051381:Rats; D011919:Rats, Inbred Strains
D011480:Protease Inhibitors; C004203:cadmium-binding protein; D002104:Cadmium; D008668:Metallothionein; D002403:Cathepsins; D019256:Cadmium Chloride