The Survival of β-lactoglobulin Peptides in the Archaeological Record: Vulnerability vs. Sequence Variation

Abstract

It is a strange observation, given the cultural co-evolution of dairying, that milk proteins are more commonly reported than any other food proteins in the archaeological record. The whey protein {beta}-lactoglobulin and in particular its eleven amino acid long peptide T125PEVDXEALEK135 seems to be preferentially preserved in both ceramic vessels and teeth (dental calculus). An amino acid substitution in the middle of the chain is valuable to track livestock management because it permits differentiation between key animal species used in dairying. The persistence of this peptide is, however, unusual as its acidic nature makes it more vulnerable to hydrolysis. Moreover, selection for the ability to digest raw milk - more specifically, the continued production of the milk sugar enzyme lactase beyond the age of normal weaning - did not begin until the early Bronze Age. It is therefore unclear why it is a milk peptide, in particular a peptide associated with the lactose-rich whey fraction, that is one of the most commonly recovered dietary peptides. The unexpected preservation of T125PEVDXEALEK135 thus presents a good case study to uncover patterns of protein survival in the archaeological record. We have previously explored the dynamics of the bovine variation of the peptide (X=Asp130) and its likelihood to undergo hydrolysis in solution. In this study, we turn our attention to the ovine (X=Asn130) and the caprine (X=Lys130) variations of the {beta}-lactoglobulin peptide to determine how the mutation in the amino acid in position 6 affects peptide conformations and vulnerability in bulk water. To do this, we use Molecular Dynamics as implemented in GROMACS 2020, with the Amber14SB forcefield and the SPC/E water model. We first perform extensive conformational analysis of both peptides in solution to determine stable structures. Then, using analogous methodology to that developed in our earlier study of the bovine peptide, we identify geometric arrangements between water and peptide that may be more prone to hydrolysis.