Home Life Sciences Degradation of apolipoprotein B-100 by lysosomal cysteine cathepsins
Article
Licensed
Unlicensed Requires Authentication

Degradation of apolipoprotein B-100 by lysosomal cysteine cathepsins

  • Martin Linke , Ronald E. Gordon , Michèle Brillard , Fabien Lecaille , Gilles Lalmanach and Dieter Brömme
Published/Copyright: September 14, 2006
Biological Chemistry
From the journal Volume 387 Issue 9

Abstract

Although the degradation of cellular or endocytosed proteins comprises the normal function of lysosomal proteinases, these enzymes were also detected extracellularly during diseases such as atherosclerosis. Since lysosomal cysteine cathepsins were demonstrated to transform native LDL particles into a proatherogenic type, the following study was undertaken to characterize the modification of LDL particles and the degradation of apolipoprotein B-100 in more detail. LDL was incubated with cathepsins B, F, K, L, S, and V at pH 5.5 and under physiological conditions (pH 7.4) for 2 h to mimic conditions of limited proteolysis. Gel electrophoretic analysis of the degradation products revealed that cathepsin-mediated proteolysis of apolipoprotein B-100 is a fast process carried out by all enzymes at pH 5.5, and by cathepsin S also at pH 7.4. Electron microscopic analysis showed that cathepsin-mediated degradation of apolipoprotein B-100 rendered LDL particles fusion-competent compared to controls. N-Terminal sequencing of cathepsin cleavage fragments from apolipoprotein B-100 revealed an abundance of enzyme-specific cleavage sites located in almost all structurally and functionally essential regions. Since the cleavage sites superimpose well with results from substrate specificity studies, they might be useful for the development of cathepsin-specific inhibitors and substrates.

Keywords: apolipoprotein B-100; cell-associated proteolysis; cleavage sites; cysteine cathepsins; LDL particle fusion; N-terminal sequencing
:
Corresponding author

References

Alves, M.F., Puzer, L., Cotrin, S.S., Juliano, M.A., Juliano, L., Bromme, D., and Carmona, A.K. (2003). S3 to S3′ subsite specificity of recombinant human Cat K and development of selective internally quenched fluorescent substrates. Biochem. J.373, 981–986.10.1042/bj20030438Search in Google Scholar

Andrews, N.W. (2000). Regulated secretion of conventional lysosomes. Trends Cell. Biol.10, 316–321.10.1016/S0962-8924(00)01794-3Search in Google Scholar

Babaev, V.R., Bobryshev, Y.V., Sukhova, G.K., and Kasantseva, I.A. (1993). Monocyte/macrophage accumulation and smooth muscle cell phenotypes in early atherosclerotic lesions of human aorta. Atherosclerosis100, 237–248.10.1016/0021-9150(93)90210-LSearch in Google Scholar

Berger, A. and Schechter, I. (1970). Mapping the active site of papain with the aid of peptide substrates and inhibitors. Philos. Trans. R. Soc. Lond. B. Biol. Sci.257, 249–264.Search in Google Scholar

Bradford, M.M. (1976). A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal. Biochem.72, 248–254.10.1016/0003-2697(76)90527-3Search in Google Scholar

Brömme, D. (2004). Cathepsin F. In: Handbook of Proteolytic Enzymes, 2nd edition, A.J. Barrett, N.D. Rawlings and J.F. Woessner, eds. (Amsterdam, The Netherlands: Elsevier Academic Press), pp. 1087–1088.Search in Google Scholar

Bromme, D., Li, Z., Barnes, M., and Mehler, E. (1999). Human cathepsin V functional expression, tissue distribution, electrostatic surface potential, enzymatic characterization, and chromosomal localization. Biochemistry38, 2377–2385.10.1021/bi982175fSearch in Google Scholar

Cardin, A.D., Witt, K.R., Chao, J., Margolius, H.S., Donaldson, V.H., and Jackson, R.L. (1984). Degradation of apolipoprotein B-100 of human plasma low density lipoproteins by tissue and plasma kallikreins. J. Biol. Chem.259, 8522–8528.10.1016/S0021-9258(17)39761-2Search in Google Scholar

Chen, G.C., Lau, K., Hamilton, R.L., and Kane, J.P. (1991). Differences in local conformation in human apolipoprotein B-100 of plasma low density and very low density lipoproteins as identified by cathepsin D. J. Biol. Chem.266, 12581–12587.10.1016/S0021-9258(18)98938-6Search in Google Scholar

Cheng, X.W., Kuzuya, M., Nakamura, K., Di, Q., Liu, Z., Sasaki, T., Kanda, S., Jin, H., Shi, G.P., Murohara, T., Yokota, M., and Iguchi, A. (2006). Localization of cysteine protease, cathepsin S, to the surface of vascular smooth muscle cells by association with integrin ανβ3. Am. J. Pathol.168, 685–694.10.2353/ajpath.2006.050295Search in Google Scholar PubMed PubMed Central

Cotrin, S.S., Puzer, L., de Souza Judice, W.A., Juliano, L., Carmona, A.K., and Juliano, M.A. (2004). Positional-scanning combinatorial libraries of fluorescence resonance energy transfer peptides to define substrate specificity of carboxydipeptidases: assays with human cathepsin B. Anal. Biochem.335, 244–252.10.1016/j.ab.2004.09.012Search in Google Scholar PubMed

De Graaf, J., Hak-Lemmers, H.L., Hectors, M.P., Demacker, P.N., Hendriks, J.C., and Stalenhoef, A.F. (1991). Enhanced susceptibility to in vitro oxidation of the dense low density lipoprotein subfraction in healthy subjects. Arterioscler. Thromb.11, 298–306.10.1161/01.ATV.11.2.298Search in Google Scholar

Fukuchi, M., Watanabe, J., Kumagai, K., Baba, S., Shinozaki, T., Miura, M., Kagaya, Y., and Shirato, K. (2002). Normal and oxidized low density lipoproteins accumulate deep in physiologically thickened intima of human coronary arteries. Lab. Invest.82, 1437–1447.10.1097/01.LAB.0000032546.01658.5DSearch in Google Scholar

Garcia-Touchard, A., Henry, T.D., Sangiorgi, G., Spagnoli, L.G., Mauriello, A., Conover, C., and Schwartz, R.S. (2005). Extracellular proteases in atherosclerosis and restenosis. Arterioscler. Thromb. Vasc. Biol.25, 1119–1127.10.1161/01.ATV.0000164311.48592.daSearch in Google Scholar

Gardella, S., Andrei, C., Lotti, L.V., Poggi, A., Torrisi, M.R., Zocchi, M.R., and Rubartelli, A. (2001). CD8+ T lymphocytes induce polarized exocytosis of secretory lysosomes by dendritic cells with release of interleukin-1β and cathepsin D. Blood98, 2152–2159.10.1182/blood.V98.7.2152Search in Google Scholar

Hakala, J.K., Oksjoki, R., Laine, P., Du, H., Grabowski, G.A., Kovanen, P.T., and Pentikainen, M.O. (2003). Lysosomal enzymes are released from cultured human macrophages, hydrolyze LDL in vitro, and are present extracellularly in human atherosclerotic lesions. Arterioscler. Thromb. Vasc. Biol.23, 1430–1436.10.1161/01.ATV.0000077207.49221.06Search in Google Scholar

Han, S.R., Momeni, A., Strach, K., Suriyaphol, P., Fenske, D., Paprotka, K., Hashimoto, S.I., Torzewski, M., Bhakdi, S., and Husmann, M. (2003). Enzymatically modified LDL induces cathepsin H in human monocytes: potential relevance in early atherogenesis. Arterioscler. Thromb. Vasc. Biol.23, 661–667.10.1161/01.ATV.0000063614.21233.BFSearch in Google Scholar

Hoff, H.F., Zyromski, N., Armstrong, D., and O'Neil, J. (1993). Aggregation as well as chemical modification of LDL during oxidation is responsible for poor processing in macrophages. J. Lipid Res.34, 1919–1929.10.1016/S0022-2275(20)35110-5Search in Google Scholar

Karpe, F. and Hamsten, A. (1994). Determination of apolipoproteins B-48 and B-100 in triglyceride-rich lipoproteins by analytical SDS-PAGE. J. Lipid Res.35, 1311–1317.10.1016/S0022-2275(20)39974-0Search in Google Scholar

Kruth, H.S. and Shekhonin, B. (1994). Evidence for loss of apo B from LDL in human atherosclerotic lesions: extracellular cholesteryl ester lipid particles lacking apo B. Atherosclerosis105, 227–234.10.1016/0021-9150(94)90053-1Search in Google Scholar

Leake, D.S. (1997). Does an acidic pH explain why low density lipoprotein is oxidised in atherosclerotic lesions? Atherosclerosis129, 149–157.10.1016/S0021-9150(96)06035-2Search in Google Scholar

Lecaille, F., Authie, E., Moreau, T., Serveau, C., Gauthier, F., and Lalmanach, G. (2001). Subsite specificity of trypanosomal cathepsin L-like cysteine proteases. Probing the S2 pocket with phenylalanine-derived amino acids. Eur. J. Biochem.268, 2733–2741.10.1046/j.1432-1327.2001.02172.xSearch in Google Scholar PubMed

Lecaille, F., Kaleta, J., and Bromme, D. (2002). Human and parasitic papain-like cysteine proteases: their role in physiology and pathology and recent developments in inhibitor design. Chem. Rev.102, 4459–4488.10.1021/cr0101656Search in Google Scholar

Lecaille, F., Weidauer, E., Juliano, M.A., Bromme, D., and Lalmanach, G. (2003). Probing cathepsin K activity with a selective substrate spanning its active site. Biochem. J.375, 307–312.10.1042/bj20030468Search in Google Scholar

Linnevers, C.J., McGrath, M.E., Armstrong, R., Mistry, F.R., Barnes, M.G., Klaus, J.L., Palmer, J.T., Katz, B.A., and Bromme, D. (1997). Expression of human cathepsin K in Pichia pastoris and preliminary crystallographic studies of an inhibitor complex. Protein Sci.6, 919–921.10.1002/pro.5560060421Search in Google Scholar

Liu, J., Sukhova, G.K., Sun, J.S., Xu, W.H., Libby, P., and Shi, G.P. (2004). Lysosomal cysteine proteases in atherosclerosis. Arterioscler. Thromb. Vasc. Biol.24, 1359–1366.10.1161/01.ATV.0000134530.27208.41Search in Google Scholar

Liu, J., Ma, L., Yang, J., Ren, A., Sun, Z., Yan, G., Sun, J., Fu, H., Xu, W., Hu, C., and Shi, G.P. (2005). Increased serum cathepsin S in patients with atherosclerosis and diabetes. Atherosclerosis186, 411–419.Search in Google Scholar

McEneny, J., Loughrey, C.M., McNamee, P.T., Trimble, E.R., and Young, I.S. (1997). Susceptibility of VLDL to oxidation in patients on regular haemodialysis. Atherosclerosis129, 215–220.10.1016/S0021-9150(96)06019-4Search in Google Scholar

McEneny, J., McMaster, C., Trimble, E.R., and Young, I.S. (2002). Rapid isolation of VLDL subfractions: assessment of composition and susceptibility to copper-mediated oxidation. J. Lipid Res.43, 824–831.10.1016/S0022-2275(20)30125-5Search in Google Scholar

Naghavi, M., John, R., Naguib, S., Siadaty, M.S., Grasu, R., Kurian, K.C., van Winkle, W.B., Soller, B., Litovsky, S., Madjid, M., et al. (2002). pH heterogeneity of human and rabbit atherosclerotic plaques; a new insight into detection of vulnerable plaque. Atherosclerosis164, 27–35.10.1016/S0021-9150(02)00018-7Search in Google Scholar

Oorni, K., Pentikainen, M.O., Ala-Korpela, M., and Kovanen, P.T. (2000). Aggregation, fusion, and vesicle formation of modified low density lipoprotein particles: molecular mechanisms and effects on matrix interactions. J. Lipid Res.41, 1703–1714.10.1016/S0022-2275(20)31964-7Search in Google Scholar

Oorni, K., Sneck, M., Bromme, D., Pentikainen, M.O., Lindstedt, K.A., Mayranpaa, M., Aitio, H., and Kovanen, P.T. (2004). Cysteine protease cathepsin F is expressed in human atherosclerotic lesions, is secreted by cultured macrophages, and modifies low density lipoprotein particles in vitro. J. Biol. Chem.279, 34776–34784.10.1074/jbc.M310814200Search in Google Scholar PubMed

Parks, W.C. (1999). Who are the proteolytic culprits in vascular disease? J. Clin. Invest.104, 1167–1168.10.1172/JCI8642Search in Google Scholar PubMed PubMed Central

Pauly, T.A., Sulea, T., Ammirati, M., Sivaraman, J., Danley, D.E., Griffor, M.C., Kamath, A.V., Wang, I.K., Laird, E.R., Seddon, A.P., et al. (2003). Specificity determinants of human cathepsin s revealed by crystal structures of complexes. Biochemistry42, 3203–3213.10.1021/bi027308iSearch in Google Scholar

Pentikainen, M.O., Oorni, K., Ala-Korpela, M., and Kovanen, P.T. (2000). Modified LDL-trigger of atherosclerosis and inflammation in the arterial intima. J. Intern. Med.247, 359–370.10.1046/j.1365-2796.2000.00655.xSearch in Google Scholar

Pillay, C.S., Elliott, E., and Dennison, C. (2002). Endolysosomal proteolysis and its regulation. Biochem. J.363, 417–429.10.1042/bj3630417Search in Google Scholar

Puzer, L., Cotrin, S.S., Alves, M.F., Egborge, T., Araujo, M.S., Juliano, M.A., Juliano, L., Bromme, D., and Carmona, A.K. (2004). Comparative substrate specificity analysis of recombinant human cathepsin V and cathepsin L. Arch. Biochem. Biophys.430, 274–283.10.1016/j.abb.2004.07.006Search in Google Scholar

Reddy, V.Y., Zhang, Q.Y., and Weiss, S.J. (1995). Pericellular mobilization of the tissue-destructive cysteine proteinases, cathepsins B, L, and S, by human monocyte-derived macrophages. Proc. Natl. Acad. Sci. USA92, 3849–3853.10.1073/pnas.92.9.3849Search in Google Scholar

Segrest, J.P., Jones, M.K., De Loof, H., and Dashti, N. (2001). Structure of apolipoprotein B-100 in low density lipoproteins. J. Lipid Res.42, 1346–1367.10.1016/S0022-2275(20)30267-4Search in Google Scholar

Simionescu, N., Vasile, E., Lupu, F., Popescu, G., and Simionescu, M. (1986). Prelesional events in atherogenesis. Accumulation of extracellular cholesterol-rich liposomes in the arterial intima and cardiac valves of the hyperlipidemic rabbit. Am. J. Pathol.123, 109–125.Search in Google Scholar

Song, J., Xu, P., Xiang, H., Su, Z., Storer, A.C., and Ni, F. (2000). The active-site residue Cys-29 is responsible for the neutral-pH inactivation and the refolding barrier of human cathepsin B. FEBS Lett.475, 157–162.10.1016/S0014-5793(00)01644-6Search in Google Scholar

Sukhova, G.K., Shi, G.P., Simon, D.I., Chapman, H.A., and Libby, P. (1998). Expression of the elastolytic cathepsins S and K in human atheroma and regulation of their production in smooth muscle cells. J. Clin. Invest.102, 576–583.10.1172/JCI181Search in Google Scholar PubMed PubMed Central

Sukhova, G.K., Zhang, Y., Pan, J.H., Wada, Y., Yamamoto, T., Naito, M., Kodama, T., Tsimikas, S., Witztum, J.L., Lu, M.L., et al. (2003). Deficiency of cathepsin S reduces atherosclerosis in LDL receptor-deficient mice. J. Clin. Invest.111, 897–906.10.1172/JCI200314915Search in Google Scholar

Torzewski, M., Suriyaphol, P., Paprotka, K., Spath, L., Ochsenhirt, V., Schmitt, A., Han, S.R., Husmann, M., Gerl, V.B., Bhakdi, S., and Lackner, K.J. (2004). Enzymatic modification of low-density lipoprotein in the arterial wall: a new role for plasmin and matrix metalloproteinases in atherogenesis. Arterioscler. Thromb. Vasc. Biol.24, 2130–2136.10.1161/01.ATV.0000144016.85221.66Search in Google Scholar PubMed

Vasile, E., Simionescu, M., and Simionescu, N. (1983). Visualization of the binding, endocytosis, and transcytosis of low-density lipoprotein in the arterial endothelium in situ. J. Cell. Biol.96, 1677–1689.10.1083/jcb.96.6.1677Search in Google Scholar

Wang, B., Shi, G.P., Yao, P.M., Li, Z., Chapman, H.A., and Bromme, D. (1998). Human cathepsin F. Molecular cloning, functional expression, tissue localization, and enzymatic characterization. J. Biol. Chem.273, 32000–32008.10.1074/jbc.273.48.32000Search in Google Scholar

Yang, C., Gu, Z.W., Yang, M., and Gotto, A.M. Jr. (1994). Primary structure of apoB-100. Chem. Phys. Lipids67–68, 99–104.10.1016/0009-3084(94)90128-7Search in Google Scholar

Yasuda, Y., Li, Z., Greenbaum, D., Bogyo, M., Weber, E., and Bromme, D. (2004). Cathepsin V, a novel and potent elastolytic activity expressed in activated macrophages. J. Biol. Chem.279, 36761–36770.10.1074/jbc.M403986200Search in Google Scholar PubMed

Zieske, A.W., Malcom, G.T., and Strong, J.P. (2002). Natural history and risk factors of atherosclerosis in children and youth: the PDAY study. Pediatr. Pathol. Mol. Med.21, 213–237.10.1080/pdp.21.2.213.237Search in Google Scholar

Published Online: 2006-09-14
Published in Print: 2006-09-01

©2006 by Walter de Gruyter Berlin New York

Articles in the same Issue

  1. The arylhydrocarbon receptor: more than a tox story
  2. The aryl hydrocarbon receptor and light
  3. The impact of aryl hydrocarbon receptor signaling on matrix metabolism: implications for development and disease
  4. A role for the aryl hydrocarbon receptor in mammary gland tumorigenesis
  5. Evidence supporting the hypothesis that one of the main functions of the aryl hydrocarbon receptor is mediation of cell stress responses
  6. The arylhydrocarbon receptor repressor (AhRR): structure, expression, and function
  7. Impact of the arylhydrocarbon receptor on eugenol- and isoeugenol-induced cell cycle arrest in human immortalized keratinocytes (HaCaT)
  8. Aryl hydrocarbon receptor agonists directly activate estrogen receptor α in MCF-7 breast cancer cells
  9. Identifying target genes of the aryl hydrocarbon receptor nuclear translocator (Arnt) using DNA microarray analysis
  10. Transcriptional signatures of immune cells in aryl hydrocarbon receptor (AHR)-proficient and AHR-deficient mice
  11. 14-3-3 proteins in membrane protein transport
  12. The K+ channel gene, Kcnb1: genomic structure and characterization of its 5′-regulatory region as part of an overlapping gene group
  13. Structure-based specificity mapping of secreted aspartic proteases of Candida parapsilosis, Candida albicans, and Candida tropicalis using peptidomimetic inhibitors and homology modeling
  14. The solution structure of the membrane-proximal cytokine receptor domain of the human interleukin-6 receptor
  15. Sequence determination of lychnin, a type 1 ribosome-inactivating protein from Lychnis chalcedonica seeds
  16. Paired helical filaments contain small amounts of cholesterol, phosphatidylcholine and sphingolipids
  17. Induction of intracellular signalling in human endothelial cells by the hyaluronan-binding protease involves two distinct pathways
  18. A novel proteolytically processed CDP/Cux isoform of 90 kDa is generated by cathepsin L
  19. Degradation of apolipoprotein B-100 by lysosomal cysteine cathepsins
  20. Identification of trypsin I as a candidate for influenza A virus and Sendai virus envelope glycoprotein processing protease in rat brain
https://doi.org/10.1515/BC.2006.160
Keywords for this article
apolipoprotein B-100; cell-associated proteolysis; cleavage sites; cysteine cathepsins; LDL particle fusion; N-terminal sequencing

Articles in the same Issue

  1. The arylhydrocarbon receptor: more than a tox story
  2. The aryl hydrocarbon receptor and light
  3. The impact of aryl hydrocarbon receptor signaling on matrix metabolism: implications for development and disease
  4. A role for the aryl hydrocarbon receptor in mammary gland tumorigenesis
  5. Evidence supporting the hypothesis that one of the main functions of the aryl hydrocarbon receptor is mediation of cell stress responses
  6. The arylhydrocarbon receptor repressor (AhRR): structure, expression, and function
  7. Impact of the arylhydrocarbon receptor on eugenol- and isoeugenol-induced cell cycle arrest in human immortalized keratinocytes (HaCaT)
  8. Aryl hydrocarbon receptor agonists directly activate estrogen receptor α in MCF-7 breast cancer cells
  9. Identifying target genes of the aryl hydrocarbon receptor nuclear translocator (Arnt) using DNA microarray analysis
  10. Transcriptional signatures of immune cells in aryl hydrocarbon receptor (AHR)-proficient and AHR-deficient mice
  11. 14-3-3 proteins in membrane protein transport
  12. The K+ channel gene, Kcnb1: genomic structure and characterization of its 5′-regulatory region as part of an overlapping gene group
  13. Structure-based specificity mapping of secreted aspartic proteases of Candida parapsilosis, Candida albicans, and Candida tropicalis using peptidomimetic inhibitors and homology modeling
  14. The solution structure of the membrane-proximal cytokine receptor domain of the human interleukin-6 receptor
  15. Sequence determination of lychnin, a type 1 ribosome-inactivating protein from Lychnis chalcedonica seeds
  16. Paired helical filaments contain small amounts of cholesterol, phosphatidylcholine and sphingolipids
  17. Induction of intracellular signalling in human endothelial cells by the hyaluronan-binding protease involves two distinct pathways
  18. A novel proteolytically processed CDP/Cux isoform of 90 kDa is generated by cathepsin L
  19. Degradation of apolipoprotein B-100 by lysosomal cysteine cathepsins
  20. Identification of trypsin I as a candidate for influenza A virus and Sendai virus envelope glycoprotein processing protease in rat brain
Sign up now to receive a 20% welcome discount
Institutional Access
How does access work?
Have an idea on how to improve our website?
Please write us.
© 2025 De Gruyter Brill
Downloaded on 13.12.2025 from https://www.degruyterbrill.com/document/doi/10.1515/BC.2006.160/html
Scroll to top button