SEQUENCE POLYMORPHISM IN KRINGLE-IV-37 IN LINKAGE DISEQUILIBRIUM WITHTHE APOLIPOPROTEIN(A) SIZE POLYMORPHISM

Apolipoprotein(a) [apo(a)] contains a variable number of identical (K- IV A/B) or nearly identical (K-IV 1, K-IV 30-37) kringle repeats that are homologous to K-IV from plasminogen. The sizes of 414 apo(a) allel es were determined by pulsed-field gel electrophoresis (PFGE) of KpnI- digested DNA. Furthermore, sequence variation in the apo(a) K-IV 30-37 domain was analysed. Reverse transcription/polymerase chain reaction (RT-PCR) cloning of human Liver poly A+ RNA followed by sequencing rev ealed a single nucleotide exchange in the ultimate K-IV (K-IV 37) of a po(a) (codon 4168); this results in an ATG (Met) to ACG (Thr) substitu tion. A PCR-based restriction assay of genomic DNA demonstrated that t his substitution represents a common polymorphism, In 231 unrelated Ty roleans, the frequencies for the K-IV 37 Thr and K-IV 37 Met alleles w ere 0.66 and 0.34, respectively. The phase between the K-IV 37 Met/Thr and the KpnI size polymorphism was determined for 224 alleles. A sign ificant linkage disequilibrium was detected between the sequence and s ize polymorphisms of apo(a). K-IV 37 Met was significantly associated with KpnI allele no. 18 (D-AB = 0.0267 +/- 0.0101; X(2) = 10.09, df = 1). The Met/Thr polymorphism was further used to test whether deletion s or duplications of K-IV 37 occur frequently in the apo(a) gene. Some 40 apo(a) alleles, 22 of which were from subjects that appeared to be double heterozygotes for K-IV repeat number and the Met/Thr variation were separated by PFGE and analysed for the 4168 Met/Thr polymorphism . The Met and Thr sequences were always present on different size alle les and no evidence for a duplication or deletion of K-IV 37 was obtai ned. This suggests that the copy number of K-IV 37 is invariable, in c ontrast to the highly variable K-IV A/B domain of the gene. The 4168 M et/Thr polymorphism had no effect on Lp(a) concentration, neither did it influence the lysine-binding property of the Lp(a) particle.