4-Hydroxy-2-nonenal-modified glyceraldehyde-3-phosphate dehydrogenase is degraded by cathepsin G.
Degradation of oxidized or oxidatively modified proteins is an essential part of the antioxidant defenses of cells. 4-Hydroxy-2-nonenal (HNE), a major reactive aldehyde formed by lipid peroxidation, causes many types of cellular damage. It has been reported that HNE-modified proteins are degraded by the ubiquitin-proteasome pathway or, in some cases, by the lysosomal pathway. However, our previous studies using U937 cells showed that HNE-modified glyceraldehyde-3-phosphate dehydrogenase (GAPDH) is degraded by an enzyme that is sensitive to a serine protease inhibitor, diisopropyl fluorophosphate (DFP), but not a proteasome inhibitor, MG-132, and that its degradation is not catalyzed in the acidic pH range where lysosomal enzymes are active. In the present study, we purified an HNE-modified GAPDH-degrading enzyme from a U937 cell extract to a final active fraction containing two proteins of 28 kDa (P28) and 27 kDa (P27) that became labeled with [(3)H]DFP. Using peptide mass fingerprinting and a specific antibody, P28 and P27 were both identified as cathepsin G. The degradation activity was inhibited by cathepsin G inhibitors. Furthermore, a cell extract from U937 cells transfected with a cathepsin G-specific siRNA hardly degraded HNE-modified GAPDH. These results suggest that cathepsin G plays a role in the degradation of HNE-modified GAPDH.
Department of Hygienic Chemistry, Showa Pharmaceutical University, 3-3165 Higashitamagawagakuen, Machida, Tokyo 194-8543, Japan.
Yukihiro
Y
Tsuchiya
Yasutaka
Y
Okuno
Kayoko
K
Hishinuma
Asami
A
Ezaki
Go
G
Okada
Mitsune
M
Yamaguchi
Toshiyuki
T
Chikuma
Hiroshi
H
Hojo
D000447:Aldehydes / Q000494:pharmacology*; D000590:Amino Acid Chloromethyl Ketones / Q000494:pharmacology; D056649:Cathepsin G; D002403:Cathepsins / Q000037:antagonists & inhibitors / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D005609:Free Radicals / Q000378:metabolism; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases / Q000737:chemistry* / Q000378:metabolism*; D006801:Humans; D010084:Oxidation-Reduction; D010449:Peptide Mapping; D034622:RNA Interference; D034741:RNA, Small Interfering / Q000235:genetics; D012697:Serine Endopeptidases / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D015842:Serine Proteinase Inhibitors / Q000494:pharmacology; D020298:U937 Cells
D000447:Aldehydes; D000590:Amino Acid Chloromethyl Ketones; D005609:Free Radicals; D034741:RNA, Small Interfering; D015842:Serine Proteinase Inhibitors; C082205:N-benzyloxycarbonylglycyl-leucyl-phenylalanine chloromethyl ketone; D005987:Glyceraldehyde-3-Phosphate Dehydrogenases; D002403:Cathepsins; D012697:Serine Endopeptidases; C534094:CTSG protein, human; D056649:Cathepsin G; C027576:4-hydroxy-2-nonenal