Mass spectral study of polymorphism of the apolipoproteins of very low density lipoprotein

Abstract

New isoforms of apolipoprotein (apo)C-I and apoC-III have been detected in delipidated fractions from very low density lipoprotein (VLDL) using matrix-assisted laser desorption (MALDI) and electrospray ionization (ESI) mass spectrometry (MS). The cleavage sites of truncated apoC-III isoforms have also been identified. The VLDL fractions were isolated by fixed-angle single-spin ultracentrifugation using a self-generating sucrose density gradient and delipidated using a newly developed C18 solid phase extraction protocol. Fifteen apoC isoforms and apoE were identified in the MALDI spectra and the existence of the more abundant species was verified by ESI-MS. The relative intensities of the apoCs are closely correlated in three normolipidemic subjects. A fourth subject with type V hyperlipidemia exhibited an elevated apoC-III level and a suppressed level of the newly discovered truncated apoC-I isoform. ApoC-II was found to be particularly sensitive to in vitro oxidation. The dynamic range and specificity of the MALDI assay shows that the complete apoC isoform profile and apoE phenotype can be obtained in a single measurement from the delipidated VLDL fraction.-Bondarenko,

Profiling of intact blood proteins by matrix‐assisted laser desorption/ionization mass spectrometry without the need for freezing – Dried serum spots as future clinical tools for patient screening

“…MALDI mass spectra generated from both (p)FFS and (p)DSS showed on average 25 intense ion signals in the mass range between m/z 6000 and 10,000 (Figure 2), resulting in good‐quality MALDI‐MS profiles for both, (p)FFS (Figure 2A) and (p)DSS preparations (Figure 2B). In all spectra, the abundant ion signals were identified as exhibiting serum proteins of adequate abundance, such as apolipoproteins, 20,28–31 complement proteins, 32 transthyretin, 5 and hemoglobin 33 (Table 2) which stands in good agreement with earlier investigations. The two most prominent ion signals were found at m/z 6631 and 6433, belonging to apolipoprotein CI (ApoCI) and a truncated apolipoprotein CI which lacked the N‐terminal dipeptide “TP” (ApoCI‐“TP”), respectively.…”

“…Treatment success with gliptins can be monitored by determining individual DPP‐IV substrate conversion rates and/or by determining surrogate substrate conversion rates. Since ApoCI is also a substrate of DPP‐IV, monitoring individual ApoCI conversion rates has been suggested to work as a surrogate marker of gliptin efficacy, 46 and MALDI‐MS has been shown in this work and by others 8,17,28 to function as an excellent readout of ApoCI truncation.…”

O-Glycoside Biomarker of Apolipoprotein C3: Responsiveness to Obesity, Bariatric Surgery, and Therapy with Metformin, to Chronic or Severe Liver Disease and to Mortality in Severe Sepsis and Graft vs Host Disease

“…22 C-terminal truncation also occurs in vivo, increasing the number of isoforms. 21,22 ApoC3 glyco-isoform ratio changes have been observed in uremia 23 and kidney disease. 24 The absolute concentration of ApoC3 is related to blood lipid content 25 and is an independent risk factor for cardiovascular disease.…”

Identification of New Apolipoprotein-CIII Glycoforms with Ultrahigh Resolution MALDI-FTICR Mass Spectrometry of Human Sera

“…Previously, MALDI-TOF and SELDI-TOF based methods have been used to study plasma/serum apolipoprotein profiles. , Although these techniques are very valuable to assign known peptides and proteins, these MS-systems lack the necessary resolution, mass accuracy and precision for a confident characterization of new species as is exemplified by earlier reports on apoCIII isoforms. , Here, the putatively identified novel apoCIII species at 9934 Da probably corresponds to the most prominent and frequently observed fucosylated apoCIII isoform (apoCIII-Hex 2 HexNAc 2 Fuc 3 ) that we characterized in the current study. The application of our ultrahigh resolution MALDI-FTICR method allowed the identification of six fucosylated apoCIII isoforms based on low-ppm precision of multiple profiles and corresponding low MME’s. , With increasing number of applications on an orbitrap system this approach is also coined as “high resolution/accurate mass (HR/AM)”.…”

“…In a few MS-based profiling studies, novel serum apoCIII glycoforms have been suggested but the exact nature of the glycan composition remained undetermined. , Further support for the presence of additional apoCIII glycoforms comes from a recent study performed in our laboratory, in which aberrantly O -glycosylated apoCIII-derived peptides were observed in urine from Schistosoma mansoni -infected individuals that were absent in noninfected individuals . Although these peptides were presumably also O -glycosylated on Thr-74 (referring to the full length protein), the glycans on these peptides were completely different from the hitherto described O -glycosylation.…”