pH-regulated formation of side products in the reductive amination approach for differential labeling of peptides in relative quantitative experiments

Among the most common stable-isotope labeling strategies, the reaction of formaldehyde with peptides in the presence of NaCNBH₃ features many attractive aspects that are conducive to its employment in quantitation experiments in proteomics. Reductive amination, with formaldehyde and d(2)-formaldehyde, is reported to be a fast, easy, and specific reaction, undoubtedly inexpensive if compared with commercially available kits for differential isotope coding. Acetaldehyde and d(4)-acetaldehyde could be employed as well without a substantial increase in terms of cost, and should provide a wider spacing between the differentially tagged peptides in the mass spectrum. Nevertheless, only a single paper reports about a diethylation approach for quantitation. We undertook a systematic analytical investigation on the reductive amination of some standard peptides pointing out the occasional occurrence of side reactions in dependence of pH or reagents order of addition, particularly observing the formation of cyclic adducts ascribable to rearrangements involving the generated Schiff-base and all the nucleophilic sites of its chemical environment. We also tried to evaluate how much this side-products amount may impair isotope coded relative quantitation

The "Bridge" Game: Role of the Fourth Player in Chiral Recognition

A new team player: The “three-point interaction” model, which is usually employed to rationalize chiral recognition, does not account for the amazing enantioselectivity measured for the receptors of many proteic acceptors. Gas-phase experiments have indicated that at least a fourth “player” must be considered: the rigidity that a receptor opposes to distortions of its cavity resulting from noncovalent interactions with a chiral molecule

Unravelling the effect of clostridia spores and lysozyme on microbiota dynamics in Grana Padano cheese : a metaproteomics approach

Grana Padano is a typical Italian Protected Designation of Origin (PDO) hard cheese largely consumed all over the world. The major problem during its production is represented by late blowing. Clostridia are gasogen bacteria responsible of the swelling during ripening, and they are partially counteracted by the use of egg white lysozyme as additive. In this work was applied, for the first time in cheese, a metaproteomic approach that identified the functional dynamics of microbial consortia in relation to the number of clostridial spores and lysozyme treatment using experimental samples of Grana Padano cheese. We used a combined custom BLAST\ua0+/MEGAN/STAMP approach to obtain a global taxonomic view associated to low and high clostridial spores cheese without and with lysozyme. Main differences were highlighted in the bacilli class. Functional analysis with SEED provided a deep view into several metabolic pathways, highlighting the subsystems \u201camino acid and derivatives\u201d and \u201cclustering-based subsystem\u201d as the targeted subsystems during lysozyme treatment in the high spore group. In these subsystems, acetate kinase from clostridia was one of the main enzymes affected by the lysozyme treatment. Biological significance Metaproteomics is a very promising and useful technique in the control of food safety and quality, from fresh products until \u2018ready to eat\u2019 food. Tools able to identify at molecular level the dynamic fingerprinting of food microbiota could be of great help to improve food safety and quality