Lipoproteomics I: Mapping of proteins in low-density lipoprotein using two-dimensional gel electrophoresis and mass spectrometry

Abstract

The molecular mechanisms underlying the relationship between low-density lipoprotein (LDL) and the risk of atherosclerosis are not clear. Therefore, detailed information about the protein composition of LDL may contribute to reveal its role in atherogenesis and the mechanisms that lead to coronary disease in humans. Here, we sought to map the proteins in human LDL by a proteomic approach. LDL was isolated by two-step discontinuous density-gradient ultracentrifugation and the proteins were separated with two-dimensional gel electrophoresis and identified with peptide mass fingerprinting, using matrix assisted laser desorption/ionization-time of flight-mass spectrometry and with amino acid sequencing using electrospray ionization tandem mass spectrometry. These procedures identified apo B-100, apo C-II, apo C-III (three isoforms), apo E (four isoforms), apo A-I (two isoforms), apo A-IV, apo J and apo M (three isoforms not previously described). In addition, three proteins that have not previously been identified in LDL were found: serum amyloid A-IV (two isoforms), calgranulin A, and lysozyme C. The identities of apo M, calgranulin A, and lysozyme C were confirmed by sequence information obtained after collision-induced dissociation fragmentation of peptides characteristic for these proteins. Moreover, the presence of lysozyme C was further corroborated by demonstrating enriched hydrolytic activity in LDL against Micrococcus lysodeikticus. These results indicate that in addition to the dominating apo B-100, LDL contains a number of other apolipoproteins, many of which occur in different isoforms. The demonstration, for the first time, that LDL contains calgranulin A and lysozyme C raises the possibility that LDL proteins may play hitherto unknown role(s) in immune and inflammatory reactions of the arterial wall.

Recent advances in blood‐related proteomics

“…Novel proteins were identified in HDL by these approaches. Also using 2-DE and the new MS technologies, Karlsson et al [100] studied the protein content of LDL, and identified calgranulin and lysozyme C in these lipoproteins. Their observation raises the possibility of new links between lipid metabolism, immunity and inflammation, which may be of relevance in the context of atherosclerosis.…”

Nanoparticle size and surface properties determine the protein corona with possible implications for biological impacts

“…The identities of the proteins attached to HDL, LDL, and VLDL have recently been analyzed using proteomics (10)(11)(12)(13)(14)(15)(16). In particular, the list of proteins believed to be associated with HDL particles has grown significantly in recent years (17).…”

Nutriproteomics: technologies and applications for identification and quantification of biomarkers and ingredients

“…Many lines of evidence strongly link them to the immune system and macrophage biology (91)(92)(93) . Several approaches have been developed for lipoprotein isolation (94)(95)(96)(97) ; delipidation (98,99) ; sample preparation (solubilisation and digestion) (100)(101)(102)(103) ; and the characterisation of protein component of lipoproteins using MS (104,105) .…”