A detailed study of specific lipoprotein parameters is necessary in individuals at high risk of coronary heart disease (CHD). During the last two decades, attention has been focused on the association of lipoprotein(a) [Lp(a)] and CHD [1,2].
Lipoprotein(a) is a cholesterol-rich lipoprotein that is an independent risk factor for the development of atherosclerosis [3]. The protein moiety of Lp(a) consists of apoli­poprotein(a) [apo(a)], joined by a disulfide linkage to apolipoprotein B-100 (apo B) [4-7]. The apo(a) gene locus on chromosome 6q26-27 (LPA) has been identified as the major quantitative trait locus (QTL) for Lp(a) concentrations [8,9]. The apo(a) gene resides within 50 kb of the plasminogen gene and shares a high degree of sequence identity with its neighbor [10].
Apolipoprotein(a) is a glycosylated protein [11] that contains a variable number of tandem repeats of a se­quence closely resembling plasminogen kringle IV (K IV VNTR), followed by a single kringle V repeat and then a region that shares 94% sequence identity with the protease domain of plasminogen [10,12]. The number of kringle IV repeats is genetically determined and is responsible for the apo(a) size heterogeneity [13] of different isoforms [14, 15]. The relative molecular mass (Mr) of the isoforms was from less than 400,000 to more than 800,000 [16]. There is an inverse trend between Lp(a) concentration and the size of its apo(a) component: low Mr isoforms (B, S1, S2) are associated with high plasma Lp(a) concentrations and high Mr isoforms (S3, S4) with intermediate Lp(a) concentrations. In the “null” phenotype no apo(a) band is detected upon immunoblotting and the Lp(a) level is low or absent [14,17]. In humans, the plasma level of Lp(a) varies from 0.1 mg/dL to over 100 mg/dL [18].
Since up to 90% of the variance in Lp(a) levels is attributable to the apo(a) locus [9,19], the possibility that polymorphisms of the apo(a) gene, other than size, could contribute to the increase of Lp(a) levels, must be considered. These include a pentanucleotide (TTTTA) repeat polymorphism in the promoter, and sequence variation in coding and non coding regions of the gene, including a C->T polymorphism at +93 (relative to the transcription start site), that creates an additional ATG start codon but also affects transcription [20-22].
We here describe a method for the separation of apo(a) isoforms and for the determination of Mr of representative apo(a) isoforms, the apo(a) allele frequencies in a Macedonian population, and the relationship between the apo(a) isoforms and plasma Lp(a) concentration.