Phenotyping apolipoprotein E*3-leiden transgenic mice by two-dimensional polyacrylamide gel electrophoresis and mass spectrometric identification.
Apolipoprotein E (ApoE) plays an important role in cholesterol and triglyceride metabolism, being one of the major structural components of chylomicrons and very low density lipoprotein (VLDL) remnants. ApoE functions as a ligand in the receptor-mediated uptake of these remnants from the blood by the liver. A variant form of ApoE, apolipoprotein E*3-Leiden, shows reduced affinity for the low density lipoprotein (LDL) receptor, and results in the dominant expression of type III hyperlipoproteinemia. Two-dimensional electrophoresis (2-DE) has been used to characterise protein expression in serum samples from control and transgenic mice expressing the human ApoE*3-Leiden mutation, fed a cholesterol-rich diet, and transgenic mice fed a normal diet. For the identification of proteins, single silver-stained spots were excised from the 2-DE gels and subjected to in-gel enzymatic digestion. Extracted peptides were analysed by matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF-MS). This proteomic approach has enabled the ApoE*3-Leiden variant to be positioned in a 2-DE separation of serum proteins, and has identified changes in the expression of haptoglobin, indicating that this protein may provide a marker for the potential onset of atherosclerosis.
Department of Analytical Sciences, SmithKline Beecham Pharmaceuticals, Harlow, Essex, UK. mark_j_skehel@sbphrd.com
J M
JM
Skehel
K
K
Schneider
N
N
Murphy
A
A
Graham
G M
GM
Benson
P
P
Cutler
P
P
Camilleri
D000595:Amino Acid Sequence; D000818:Animals; D053318:Apolipoprotein E3; D001057:Apolipoproteins E / Q000097:blood* / Q000145:classification / Q000235:genetics; D002791:Cholesterol, Dietary / Q000378:metabolism; D015180:Electrophoresis, Gel, Two-Dimensional / Q000379:methods; D006801:Humans; D051379:Mice; D008822:Mice, Transgenic; D008969:Molecular Sequence Data; D010641:Phenotype; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization / Q000379:methods
D053318:Apolipoprotein E3; D001057:Apolipoproteins E; D002791:Cholesterol, Dietary; C052720:apolipoprotein E3 (Leidein)