No abstract

Chemical communication in elephants has been well studied at the chemical and behavioural levels. Pheromones have been identified in the Asian elephant (Elephas maximus), including (Z)-7-dodecenyl acetate and frontalin, and their specific effects on the sexual behaviour of elephants have been accurately documented. In contrast, our knowledge on the proteins mediating detection of pheromones in elephants remains poor and superficial, with only three annotated and reliable entries in sequence databases, two of them being odorant-binding proteins (OBPs), and the third a member of von Ebner's gland (VEG) proteins. Proteomic analysis of trunk wash extract from African elephant (Loxodonta africana) identified one of the OBPs (LafrOBP1) as the main component. We therefore expressed LafrOBP1 and its Asian elephant orthologue in yeast Pichia pastoris and found that both recombinant proteins, as well as the natural LafrOBP1 are tuned to (Z)-7-dodecenyl acetate, but have no affinity for frontalin. Both the natural and recombinant LafrOBP1 carry post-translational modifications such as O-glycosylation, phosphorylation and acetylation, but as these modifications affect only a very small amount of the protein, we cannot establish their potential effects on the ligand-binding properties of OBP1.

To investigate thiol-disulfide interchange reactions in heated milk yielding non-native intramolecular rearranged and intermolecular cross-linked proteins, a proteomic study based on nanoLC-ESI-Q-Orbitrap-MS/MS and dedicated bioinformatics was accomplished. Raw milk samples heated for different times and various commercial dairy products were analyzed. Qualitative experiments on tryptic digests of resolved protein mixtures assigned the corresponding disulfide-linked peptides. Results confirmed the limited data available on few milk proteins, generated the widest inventory of components (63 in number) involved in thioldisulfide exchange processes, and provided novel structural information on S−S-bridged molecules. Quantitative experiments on unresolved protein mixtures from both sample typologies estimated the population of molecules associated with thiol-disulfide reshuffling processes. Disulfide-linked peptides associated with native intramolecular S−S bonds generally showed a progressive reduction depending on heating time/harshness, whereas those related to specific non-native intramolecular/intermolecular ones showed an opposite quantitative trend. This was associated with a temperature-dependent augmented reactivity of definite native protein thiols and S−S bridges, which determined the formation of non-native rearranged monomers and cross-linked oligomers. Results provided novel information for possibly linking the nature and extent of thiol-disulfide exchange reactions in heated milk proteins to the corresponding functional and technological characteristics, with possible implications on food digestibility, allergenicity, and bioactivity.