Mass spectral analysis of the apolipoproteins on dog (Canis lupus familiaris) high density lipoproteins. Detection of apolipoprotein A-II

In a recent study, we reported the detection of apoA-II associated with the plasma high density lipoproteins of pigs that were previously thought to lack or to have this apolipoprotein in trace amounts. Dogs have also been reported to lack this apolipoprotein; however, genomic data have revealed that the gene for apoA-II is present on chromosome 38. Prompted by this finding, we have carried out detailed mass spectral measurements on dog apo HDL. The molecular mass of apoA-II was obtained as well as values for proapoA-I, apoA-I, apoC-I. In each case, the measured values were found to be in excellent agreement with the corresponding molecular weights calculated from genomic data. Following reverse-phase chromatography, tryptic fragments in selected fractions were analyzed by tandem mass spectrometry (MSMS). In addition to apoA-I, proapoA-I and apoA-II, enzymatic fragments from both apoC-II and apoA-IV were detected. Post-translational modification (PTM) of apoA-I, involving glycosylation, oxidation of a single methionine and acylation, were also noted. We also report on the sequencing of apoC-I using “Top Down” mass spectrometry analysis.