Isolation and characterization of four heparin-binding cyanogen bromide peptides of human plasma apolipoprotein B

Publisher Website
Google Scholar
Abstract
Apolipoprotein B-100 (apoB-100) is the major protein constituent of human plasma low-density lipoproteins (LDL). On the basis of its amino acid sequence [Chen, S.-H., Yang, C.-Y., Chen, P.-F., Setzer, D., Tanimura, M., Li, W.-H., Gotto, A. M., Jr., and Chan, L. (1986) J. Biol. Chem. 261, 12918-12921], apo B-100 is one of the largest monomeric proteins known with a calculated molecular weight of 512,937. Heparin binds to the LDL surface by interacting with positively charged amino acid residues of apoB-100, forming soluble complexes in the absence of divalent metals and insoluble complexes in their present. The purpose of this study was to isolate and characterize the heparin-binding domain(s) of apoB-100. Human plasma LDL were fragmented with cyanogen bromide (CNBr). After delipidation and reduction-carboxymethylation, the CNBr peptides were fractionated by sequential chromatography on DEAE-Sephacel, Mono S, and high reactive heparin (HRH) AffiGel-10; HRH was purified by chromatography of crude bovine lung heparin on LDL AffiGel-10. Heparin-binding peptides were further purified by reverse-phase high-performance liquid chromatography. Heparin-binding activity was monitored by a dot-blot assay with 125I-HRH. The amino-terminal sequences of four CNBr heparin-binding peptides (CNBr-I-IV) were determined. CNBr-I-IV correspond to residues 2016-2151, 3109-3240, 3308-3394, and 3570-3719, respectively, of the amino sequence of apoB-100. Each CNBr peptide contains a domain(s) of basic amino acid residues which we suggest accounts for their heparin-binding activity. CNBr-I is located near the middle of the apoB-100 sequence whereas CNBr-II,-III, and -IV are clustered near the carboxyl-terminal end of the protein. The proposed heparin-binding domains of CNBr-II (residues 3150-3157) and -IV (residues 3670-3677) show structural homology to the reported heparin-binding region of human vitronectin; CNBR-III (residues 3361-3368) shows structural homology to the known receptor/heparin-binding region of apolipoprotein E (residues 144-151). We suggest that one or more of these heparin-binding domains possibly corresponds to the receptor-binding region(s) of LDL as heparin is known to displace LDL from its membrane receptor.

This publication has 46 references indexed in Scilit: