Detection of a single nucleotide polymorphism in the human alpha-lactalbumin gene: implications for human milk proteins.

Variability in the protein composition of breast milk has been observed in many women and is believed to be due to natural variation of the human population. Single nucleotide polymorphisms (SNPs) are present throughout the entire human genome, but the impact of this variation on human milk composition and biological activity and infant nutrition and health is unclear. The goals of this study were to characterize a variant of human alpha-lactalbumin observed in milk from a Filipino population by determining the location of the polymorphism in the amino acid and genomic sequences of alpha-lactalbumin. Milk and blood samples were collected from 20 Filipino women, and milk samples were collected from an additional 450 women from nine different countries. alpha-Lactalbumin concentration was measured by high-performance liquid chromatography (HPLC), and milk samples containing the variant form of the protein were identified with both HPLC and mass spectrometry (MS). The molecular weight of the variant form was measured by MS, and the location of the polymorphism was narrowed down by protein reduction, alkylation and trypsin digestion. Genomic DNA was isolated from whole blood, and the polymorphism location and subject genotype were determined by amplifying the entire coding sequence of human alpha-lactalbumin by PCR, followed by DNA sequencing. A variant form of alpha-lactalbumin was observed in HPLC chromatograms, and the difference in molecular weight was determined by MS (wild type=14,070 Da, variant=14,056 Da). Protein reduction and digestion narrowed the polymorphism between the 33rd and 77th amino acid of the protein. The genetic polymorphism was identified as adenine to guanine, which translates to a substitution from isoleucine to valine at amino acid 46. The frequency of variation was higher in milk from China, Japan and Philippines, which suggests that this polymorphism is most prevalent in Asia. There are SNPs in the genome for human milk proteins and their implications for protein bioactivity and infant nutrition need to be considered.

A multinational study of alpha-lactalbumin concentrations in human milk.

Alpha-lactalbumin, a 14-kD protein, plays a central biochemical role in the mammary gland as the regulatory subunit of lactose synthase, and also plays a nutritional role for the rapidly growing neonate as the protein in highest concentration in human milk. The current study was undertaken to better characterize alpha-lactalbumin concentrations in human milk from a variety of countries. Mature human milk (lactation duration > or =1 month) was collected from at least 50 women from nine different countries on five continents. Alpha-lactalbumin concentration was determined by HPLC. The mean +/- SD for 452 samples was 2.44 +/- 0.64 g/L. The mean value of the samples from the United States was significantly higher than that from any other country, and the mean in Mexico was significantly lower than that from every country except China and Canada. Alpha-lactalbumin concentration decreased with increasing duration of lactation and was positively correlated with total nitrogen. On average, alpha-lactalbumin contributed 16% of the total nitrogen content of human milk and consequently an important part of the amino acid content.