Metabolism and degradation products of recombinant human insulin-like growth factor-I in lysosomes of rat kidney.
1. The degradation of recombinant human insulin-like growth factor-I (rhIGF-I) by purified lysosomes of rat kidney was examined in vitro. The peptide structures of the 13 degradation products were deduced from the sequence analysis and the molecular mass. Rat kidney lysosomal cathepsins efficiently cleave rhIGF-I to two chain peptides, like insulin. The cleavages mainly occur at the C-peptide/A-chain junction, D-peptide/A-chain junction and B21-22 or B22-23. 2. The effect of inhibitors on the lysosomal degradation of rhIGF-I was examined semiquantitatively by the rate of formation of the degradation products. The degradation of rhIGF-I was almost completely inhibited by the lysosomal cysteine protease inhibitors, leupeptin and leucine chloromethyl ketone, and a serine protease inhibitor, phenylmethylsulphonyl fluoride. On the other hand, the degradation was enhanced by the addition of a reducing agent, glutathione.
Tanaka
Y
Y
Biopharmaceutical and Pharmacokinetic Research Laboratories, Fujisawa Pharmaceutical Co., Ltd, Osaka, Japan.
Tamoto
H
H
Tozuka
Z
Z
Sato
A
A
Kimura
T
T
D000590:Amino Acid Chloromethyl Ketones / Q000494:pharmacology; D000595:Amino Acid Sequence; D000818:Animals; D002851:Chromatography, High Pressure Liquid / Q000379:methods; D015853:Cysteine Proteinase Inhibitors / Q000494:pharmacology; D005978:Glutathione / Q000378:metabolism / Q000494:pharmacology; D006801:Humans; D007328:Insulin / Q000378:metabolism; D007334:Insulin-Like Growth Factor I / Q000235:genetics / Q000378:metabolism* / Q000493:pharmacokinetics; D007668:Kidney / Q000187:drug effects / Q000378:metabolism*; D007976:Leupeptins / Q000378:metabolism / Q000494:pharmacology; D008247:Lysosomes / Q000187:drug effects / Q000378:metabolism*; D008297:Male; D008854:Microscopy, Electron; D008969:Molecular Sequence Data; D010446:Peptide Fragments / Q000032:analysis / Q000378:metabolism; D011384:Proinsulin / Q000378:metabolism; D051381:Rats; D017207:Rats, Sprague-Dawley; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism / Q000493:pharmacokinetics; D017421:Sequence Analysis
D000590:Amino Acid Chloromethyl Ketones; D015853:Cysteine Proteinase Inhibitors; D007328:Insulin; D007976:Leupeptins; D010446:Peptide Fragments; D011994:Recombinant Proteins; C026937:leucine chloromethyl ketone; D007334:Insulin-Like Growth Factor I; D011384:Proinsulin; D005978:Glutathione; C032854:leupeptin