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Journal of Lipid Research, Vol 32, 1529-1535, Copyright © 1991 by Lipid Research, Inc.
JE Hixson and PK Powers
Genetic polymorphisms of apolipoprotein A-IV (apoA-IV) have been detected
by isoelectric focusing of serum proteins. Because genetic variation in
apoA-IV has significant effects on lipid risk factors, we used restriction
enzyme isoform genotyping (restriction isotyping) to determine apoA-IV
isoform genotypes at the DNA level for a large population (n = 509). In
contrast to isoelectric focusing methods, restriction isotyping relies on
nucleotide differences, enabling unambiguous typing of known isoforms and
detection of new alleles that mimic other isoforms with shared charge
properties. To determine genotypes for the common A-IV-1 isoform (Gln at aa
position 360) and A- IV-2 isoform (360His), we used a mismatched primer for
polymerase chain reaction (PCR) to introduce a restriction site (PvuII)
that distinguishes each isoform. Using a portion of the same PCR reaction,
we used HinfI to distinguish isoforms with Thr at position 347 (347Thr)
versus Ser (347Ser). In surveys for these common genotypes, we detected
heterozygotes for an allele with an insertion of 12 bp. Nucleotide
sequencing showed that this allele is identical to the A-IV-0 isoform that
inserts a hydrophilic repeat (Glu Gln Gln Gln) in a conserved region near
the carboxy terminus. In addition, we discovered a new allele with a 12 bp
deletion that removes a repeat (Glu Gln Gln Gln) from the same region.
Nucleotide sequencing showed that this allele removes an acidic charge
relative to A-IV-1, so we have named this isoform A-IV-2*. This isoform has
not been discovered at the protein level, perhaps due to shared charge
properties with A-IV-2 isoforms.
ARTICLES
Restriction isotyping of human apolipoprotein A-IV: rapid typing of known isoforms and detection of a new isoform that deletes a conserved repeat
Department of Genetics, Southwest Foundation for Biomedical Research, San Antonio, TX 78228-0147.
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