Heterogeneity of apolipoprotein B: isolation of a new species from human chylomicrons.

Abstract

Low density lipoproteins and the triglyceride-rich lipoproteins of human serum each contain proteins of high molecular weight termed apolipoprotein B,which have previously been thought to be identical. We have isolated four species of apolipoprotein B with unique molecular weights and amino acid compositions. We have assigned numerical designations to these species in a centile system based upon their relative apparent Mr in NaDodSO4. One which we term B-100, with an apparent Mr of 549,000 ± 7650 (SD) determined by NaDodSO4 gel electrophoresis, predominates in low density and very low density lipoproteins and is also present in chylomicrons from thoracic duct lymph or from plasma. Substantial amounts of two large proteins designated B-74 (apparent Mr 407,000 ± 5790) and B-26 (apparent Mr 144,500 + 8970), which appear to be complementary fragments or constituents of the B-100 protein, are found in the low density lipoproteins of many individuals. A distinct protein, B-48, with an apparent Mr of 264,000 i 8150 is a major and constant constituent of chylomicrons from thoracic duct lymph or from plasma.The lipoproteins of human serum contain at least 12 species of proteins' (apolipoproteins) (1). The most abundant of these, apolipoprotein B (apo B) shares with another species, apolipoprotein E, the ability to interact with high-affinity receptors on cell membranes (2). apo B is distinguished by its apparent inability to transfer among lipoprotein particles, by its extreme insolubility in aqueous buffers after the lipid moiety is removed, and by its insolubility in 4.2 M tetramethylurea (3). Studies of apo B solubilized with various denaturing solvents and amphiphilic agents have yielded widely varying estimates of molecular weight (1, 4-7).We describe here the presence of four major molecular species of apo B. One of these is nearly the sole form in very low density lipoproteins (VLDL). It also appears in low density lipoproteins (LDL) and chylomicrons from thoracic duct lymph and plasma. Two others appear to be fragments or components of the former and appear only in LDL. The fourth, distinguished by both its molecular weight and amino acid composition, is found only in chylomicrons.MATERIALS AND METHODS Preparation of Lipoproteins. Serum was prepared from blood obtained from donors who had fasted for 12-16 hr. Lipoproteins of serum were separated by repetitive preparative ultracentrifugation in a 40.3 rotor, at 12°C, in a Beckman model L ultracentrifuge (8). VLDL were separated in the p < 1.006 g/cm3 fraction of serum from normolipidemic individuals after centrifugation for 18 hr. The ultracentrifugation was repeated once. Chylomicrons were judged to be absent from preparations of VLDL by the absence of a visible supernatant layer after refrigeration for 24 hr at 40C and by the absence of lipoproteinsThe publication costs of this article were defrayed in part by page charge payment. This article must therefore be hereby marked "ad- Measures to Prevent Nonspecific Degradation of apo B. Gentamycin sulfate, ...

Measurement of very low density and low density lipoprotein apolipoprotein (Apo) B-100 and high density lipoprotein Apo A-I production in human subjects using deuterated leucine. Effect of fasting and feeding.

“…A stable isotope method for measuring VLDL apo B synthesis (more correctly termed production) has recently been published by Cryer et al (5 (14,25). Second, we used the plateau enrichment of VLDL apo B-100 to estimate the enrichment of the tissue amino acid precursor pool, instead of the plateau enrichment of urinary hippurate.…”

Glycated LDL Concentrations in Non-Diabetic and Diabetic Subjects Measured with Monoclonal Antibodies Reactive with Glycated Apolipoprotein B Epitopes

“…Apolipoprotein B is a complex macromolecule that exists in two primary forms in human plasma (30,31), and it is the protein determinant for the cellular recognition and uptake of LDL by the LDL receptor (32)(33)(34)(35). The binding of apolipoprotein B to the LDL receptor results in internalization and degradation of LDL, promoting the clearance of LDL from the plasma and regulating intracellular cholesterol handling and biosynthesis (36)(37)(38).…”

Evidence for a structural relationship between apoB75 kDa and human plasma apolipoprotein B100, from translation of human liver mRNA in vitro and immunochemical studies with monoclonal and polyclonal antibodies

“…The possibility that this protein has a subunit structure has been suggested by the observations of several authors [I -10,271. It has however been argued that apoB proteins with lower molecular mass than apoBlOO are proteolytic fragments formed in vivo [l] or during the isolation procedure [1,25] either by proteases [28] or by the influence of autooxidizing lipids [26,29].…”