Polymorphism at donkey β-lactoglobulin II locus: identification and characterization of a new genetic variant with a very low expression

Andrea Criscione, Vincenzo Cunsolo, Serena Tumino, Antonella Di Francesco, Salvatore Bordonaro, Vera Muccilli, Rosaria Saletti, Donata Marletta

In the last years, donkey milk had evidenced a renewed interest as a potential functional food and a breast milk substitute. In this light, the study of the protein composition assumes an important role. In particular, β-lactoglobulin (β-LG), which is considered as one of the main allergenic milk protein, in donkey species consists of two molecular forms, namely β-LG I and β-LG II. In the present research, a genetic analysis coupled with a proteomic approach showed the presence of a new allele, here named F, which is apparently associated with a null or a severely reduced expression of β-LG II protein. The new β-LG II F genetic variant shows a theoretical average mass (Mav) of 18,310.64 Da, a value practically corresponding with that of the variant D (∆mass < 0.07 Da), but differs from β-LG II D for two amino acid substitutions: Thr100(variant F) → Ala100(variant D) and Thr118(variant F) → Met118(variant D). Proteomic investigation of the whey protein fraction of an individual milk sample, homozygous FF at β-LG II locus, allowed to identify, as very minor component, the new β-LG II F genetic variant. By MS/MS analysis of enzymatic digests, the sequence of the β-LG II F was characterized, and the predicted genomic data confirmed.