Arteriosclerosis, Vol 9, 96-108, Copyright © 1989 by American Heart Association

ARTICLES

Structure of apolipoprotein B-100 of human low density lipoproteins

CY Yang, ZW Gu, SA Weng, TW Kim, SH Chen, HJ Pownall, PM Sharp, SW Liu, WH Li and AM Gotto Jr
Department of Medicine, Baylor College of Medicine, Houston, Texas.

We have analyzed low density lipoproteins (LDL) apolipoprotein (apop) B structure by direct sequence analysis of LDL apo B-100 tryptic peptides. Native LDL were digested with trypsin, and the products were fractionated on a Sephadex G-50 column. The partially digested apo B- 100 still associated with lipids was recovered in the void volume (designated trypsin-nonreleasable, TN, peptides). The released peptides (designated trypsin-releasable, TR, peptides) in subsequent peaks were repurified on two successive high-performance liquid chromatography (HPLC) columns. The TN peak was delipidated and redigested with trypsin, and the resulting peptides were purified on two successive HPLC columns. Using this approach, we sequenced over 88% of LDL apo B- 100, extending and refining our previous study (Nature 1986;323:738- 742) which covered 52% of the protein. TN peptides made up 31%, and the TR peptides, 34% of the apo B-100 sequence; 23.7% were found under both TN and TR categories. Based on its differential trypsin releasability, apo B-100 can be divided into five domains: 1) residues 1----1000, largely TR; 2) residues 1001----1700, alternating TR and TN; 3) residues 1701----3070, largely TN; 4) residues 3071----4100, mainly TR and mixed; and 5) residues 4101----4536, almost exclusively TN. Domain 1 contained 14 of the 25 Cys residues in apo B. Domain 4 encompassed seven N-glycosylation sites, and contained the putative receptor binding domains. All 19 potential N-glycosylation sites were directly sequenced: 16 were found to be glycosylated and three were not. Three pairs of disulfide bridges were also mapped. Finally, a combination of cDNA sequencing, direct mRNA sequencing, and comparison of published apo B-100 sequences allowed us to identify specific amino acid residues within apo B-100 that seem to represent bona fide allelic variations. Our study provides information on LDL apo B-100 structure that will be important to our understanding of its conformation and metabolism.

This article has been cited by other articles:

				L. Wang, M. T. Walsh, and D. M. Small Apolipoprotein B is conformationally flexible but anchored at a triolein/water interface: A possible model for lipoprotein surfaces PNAS, May 2, 2006; 103(18): 6871 - 6876. [Abstract] [Full Text] [PDF]

				A. S. Ledford, R. B. Weinberg, V. R. Cook, R. R. Hantgan, and G. S. Shelness Self-association and Lipid Binding Properties of the Lipoprotein Initiating Domain of Apolipoprotein B J. Biol. Chem., March 31, 2006; 281(13): 8871 - 8876. [Abstract] [Full Text] [PDF]

				A. Harazono, N. Kawasaki, T. Kawanishi, and T. Hayakawa Site-specific glycosylation analysis of human apolipoprotein B100 using LC/ESI MS/MS Glycobiology, May 1, 2005; 15(5): 447 - 462. [Abstract] [Full Text] [PDF]

				A. J. Whitfield, P. H. R. Barrett, F. M. van Bockxmeer, and J. R. Burnett Lipid Disorders and Mutations in the APOB Gene Clin. Chem., October 1, 2004; 50(10): 1725 - 1732. [Abstract] [Full Text] [PDF]

				G. N. Fredrikson, B. Hedblad, G. Berglund, R. Alm, M. Ares, B. Cercek, K.-Y. Chyu, P. K. Shah, and J. Nilsson Identification of Immune Responses Against Aldehyde-Modified Peptide Sequences in ApoB Associated With Cardiovascular Disease Arterioscler. Thromb. Vasc. Biol., May 1, 2003; 23(5): 872 - 878. [Abstract] [Full Text] [PDF]

				X. Wang, V. Chauhan, A. T. Nguyen, J. Schultz, J. Davignon, S. G. Young, J. Boren, T. L. Innerarity, H. Rutai, and R. W. Milne Immunochemical evidence that human apoB differs when expressed in rodent versus human cells J. Lipid Res., March 1, 2003; 44(3): 547 - 553. [Abstract] [Full Text] [PDF]

				C. Pallaud, R. Gueguen, C. Sass, M. Grow, S. Cheng, G. Siest, and S. Visvikis Genetic influences on lipid metabolism trait variability within the Stanislas Cohort J. Lipid Res., November 1, 2001; 42(11): 1879 - 1890. [Abstract] [Full Text] [PDF]

				J. P. Segrest, M. K. Jones, H. De Loof, and N. Dashti Structure of apolipoprotein B-100 in low density lipoproteins J. Lipid Res., September 1, 2001; 42(9): 1346 - 1367. [Abstract] [Full Text] [PDF]

				J. A. DeLozier, J. S. Parks, and G. S. Shelness Vesicle-binding properties of wild-type and cysteine mutant forms of {{alpha}}1 domain of apolipoprotein B J. Lipid Res., March 1, 2001; 42(3): 399 - 406. [Abstract] [Full Text]

				H. Herscovitz, A. Derksen, M. T. Walsh, C. J. McKnight, D. L. Gantz, M. Hadzopoulou-Cladaras, V. Zannis, C. Curry, and D. M. Small The N-terminal 17% of apoB binds tightly and irreversibly to emulsions modeling nascent very low density lipoproteins J. Lipid Res., January 1, 2001; 42(1): 51 - 59. [Abstract] [Full Text]

				D. L. Gantz, M. T. Walsh, and D. M. Small Morphology of sodium deoxycholate-solubilized apolipoprotein B-100 using negative stain and vitreous ice electron microscopy J. Lipid Res., September 1, 2000; 41(9): 1464 - 1472. [Abstract] [Full Text]

				X. Wang, R. Pease, J. Bertinato, and R. W. Milne Well-Defined Regions of Apolipoprotein B-100 Undergo Conformational Change During Its Intravascular Metabolism Arterioscler. Thromb. Vasc. Biol., May 1, 2000; 20(5): 1301 - 1308. [Abstract] [Full Text] [PDF]

				X. F. Huang and G. S. Shelness Efficient glycosylation site utilization by intracellular apolipoprotein B: implications for proteasomal degradation J. Lipid Res., December 1, 1999; 40(12): 2212 - 2222. [Abstract] [Full Text] [PDF]

				C.-y. Yang, Z.-W. Gu, M. Yang, S.-N. Lin, A. J. Garcia-Prats, L. K. Rogers, S. E. Welty, and C. V. Smith Selective modification of apoB-100 in the oxidation of low density lipoproteins by myeloperoxidase in vitro J. Lipid Res., April 1, 1999; 40(4): 686 - 698. [Abstract] [Full Text]

				G. S. Shelness, M. F. Ingram, X. F. Huang, and J. A. DeLozier Apolipoprotein B in the Rough Endoplasmic Reticulum: Translation, Translocation and the Initiation of Lipoprotein Assembly J. Nutr., February 1, 1999; 129(2): 456 - 456. [Abstract] [Full Text] [PDF]

				E. Nicodeme, F. Benoist, R. McLeod, Z. Yao, J. Scott, C. C. Shoulders, and T. Grand-Perret Identification of Domains in Apolipoprotein B100 That Confer a High Requirement for the Microsomal Triglyceride Transfer Protein J. Biol. Chem., January 22, 1999; 274(4): 1986 - 1993. [Abstract] [Full Text] [PDF]

				W. Liao, S.-C. J. Yeung, and L. Chan Proteasome-mediated Degradation of Apolipoprotein B Targets Both Nascent Peptides Cotranslationally Before Translocation and Full-length Apolipoprotein B After Translocation into the Endoplasmic Reticulum J. Biol. Chem., October 16, 1998; 273(42): 27225 - 27230. [Abstract] [Full Text] [PDF]

				J. P. Segrest, M. K. Jones, V. K. Mishra, V. Pierotti, S. H. Young, J. Borén, T. L. Innerarity, and N. Dashti Apolipoprotein B-100: conservation of lipid-associating amphipathic secondary structural motifs in nine species of vertebrates J. Lipid Res., January 1, 1998; 39(1): 85 - 102. [Abstract] [Full Text]

				F. Benoist and T. Grand-Perret Co-translational Degradation of Apolipoprotein B100 by the Proteasome Is Prevented by Microsomal Triglyceride Transfer Protein. SYNCHRONIZED TRANSLATION STUDIES ON HepG2 CELLS TREATED WITH AN INHIBITOR OF MICROSOMAL TRIGLYCERIDE TRANSFER PROTEIN J. Biol. Chem., August 15, 1997; 272(33): 20435 - 20442. [Abstract] [Full Text] [PDF]

				M. F. Ingram and G. S. Shelness Folding of the Amino-terminal Domain of Apolipoprotein B Initiates Microsomal Triglyceride Transfer Protein-dependent Lipid Transfer to Nascent Very Low Density Lipoprotein J. Biol. Chem., April 11, 1997; 272(15): 10279 - 10286. [Abstract] [Full Text] [PDF]

				M. S. Bolgar, C.-Y. Yang, and S. J. Gaskell First Direct Evidence for Lipid/Protein Conjugation in Oxidized Human Low Density Lipoprotein J. Biol. Chem., November 8, 1996; 271(45): 27999 - 28001. [Abstract] [Full Text] [PDF]

				P. Sivaram, T. Vanni-Reyes, and I. J. Goldberg Endothelial Cells Synthesize and Process Apolipoprotein B J. Biol. Chem., June 21, 1996; 271(25): 15261 - 15266. [Abstract] [Full Text] [PDF]

				D. G. Gretch, S. L. Sturley, L. Wang, B. A. Lipton, A. Dunning, K. A. A. Grunwald, J. R. Wetterau, Z. Yao, P. Talmud, and A. D. Attie The Amino Terminus of Apolipoprotein B Is Necessary but Not Sufficient for Microsomal Triglyceride Transfer Protein Responsiveness J. Biol. Chem., April 12, 1996; 271(15): 8682 - 8691. [Abstract] [Full Text] [PDF]

				R. J. Pease, J. M. Leiper, G. B. Harrison, and J. Scott Studies on the Translocation of the Amino Terminus of Apolipoprotein B into the Endoplasmic Reticulum J. Biol. Chem., March 31, 1995; 270(13): 7261 - 7271. [Abstract] [Full Text] [PDF]