Leucyl-lysine

Glutathione-rich inactivated dry yeasts (GSH-IDY) are purported to accumulate glutathione intracellularly and then released into the must. Glutathione is beneficial for wine quality, but research has highlighted that GSH-IDYs have a synergic antioxidant effect similar to that of molecular GSH. Combination of negative mode ultra-high-resolution Fourrier-Transform Ion-Cyclotron-Resonance Mass Spectrometry ((-)FT-ICR-MS), ultra-high-performance liquid chromatography coupled to a Quadrupole-Time of Flight mass spectrometer (UHPLC-Q-ToF-MS) and HPLC/Diode Detector Array (DAD)-Fluorescence spectroscopy was applied to three inactivated dry yeasts soluble fractions, with increasing intracellular glutathione concentration, in order to explore the chemical diversity released in different synthetic media. Using the mean of size exclusion chromatography/DAD and fluorescence detection we report than most of the signals detected were below the 5-75 kDa-calibrated region of the chromatogram, indicating that most of the soluble protein fraction is composed of low molecular weight soluble peptides. In light of these results, high-resolution mass spectrometry was used to scan and annotate the low molecular weight compounds from 50 to 1500 Da and showed that GSH level of enrichment in IDYs was correlated to a discriminant chemical diversity of the corresponding soluble fractions. Our results clearly show an impact of the GSH accumulation process not only visible on the glutathione itself, but also on the global diversity of compounds. Within the 1674 ions detected by (-)FT-ICR-MS, the ratio of annotated elemental formulas containing carbon, hydrogen, oxygen, nitrogen and sulfur (CHONS) to annotated elemental formulas containing carbon, hydrogen, oxygen (CHO) increased from 0.2 to 2.1 with the increasing levels of IDYs GSH content and 36 unique CHONS annotated formulas were unique to the IDY with the highest concentration of GSH. Amongst the 1674 detected ions 193 were annotated as potential peptides (from 2 to 5 residues), 61 ions were annotated as unique amino acid combinations and 46% of which being significantly more intense in GSH-rich IDY. Thus, the process leading to the accumulation of glutathione also involves other metabolic pathways which contribute to an increase in CHONS containing compounds potentially released in wine, notably peptides.

In this paper, some cationic dipeptides from scallop (Patinopecten yessoensis) male gonads (SMGs), which can synergistically gel with ι-carrageenan (ι-C), were screened by the in silico approach. Fourteen protein sequences of SMGs were obtained by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) and nano liquid chromatography-mass spectrometry/mass spectrometry (nanoLC-MS/MS) analysis and were then hydrolyzed via in silico simulation. A total of 414 sequences were obtained with 56 duplicates, half of which were positively charged at pH 7. Among the cation sequences, 171 had good water solubility, including two amino acids (Lys and Arg). The molecular weight analysis of the cationic water-soluble sequences showed that 0.2-0.3 kDa accounted for the highest proportion. Based on the obvious synergistic effect of Lys and ι-C, 11 Lys-containing dipeptides, including Ser-Lys (SK), Thr-Lys (TK), Trp-Lys (WK), Ala-Lys (AK), Leu-Lys (LK), Gly-Lys (GK), Val-Lys (VK), Cys-Lys (CK), Asn-Lys (NK), Phe-Lys (FK), and Met-Lys (MK), were finally screened out to study gelation with ι-C. It was found that the dipeptides/ι-C formed firm gels except WK/ι-C. The values of the storage modulus (G') of 11 dipeptides/ι-C were investigated by a rheometer. The G' of 8 dipeptides/ι-C was higher than 1000 Pa. These results indicated that the in silico-screened dipeptides from SMGs can form composite gels with ι-C, which can be used for the design and development of functional hydrogels.

In different bacteria, primarily cytosolic and metabolic proteins are characterized as surface localized and interacting with different host factors. These moonlighting proteins include glycolytic enzymes, and it has been hypothesized that they influence the virulence of pathogenic species. The presence of surface-displayed glycolytic enzymes and their interaction with human plasminogen as an important host factor were investigated in the genome-reduced and cell wall-less microorganism Mycoplasma pneumoniae, a common agent of respiratory tract infections of humans. After successful expression of 19 glycolytic enzymes and production of polyclonal antisera, the localization of proteins in the mycoplasma cell was characterized using fractionation of total proteins, colony blot, mild proteolysis and immunofluorescence of M. pneumoniae cells. Eight glycolytic enzymes, pyruvate dehydrogenases A to C (PdhA-C), glyceraldehyde-3-phosphate dehydrogenase (GapA), lactate dehydrogenase (Ldh), phosphoglycerate mutase (Pgm), pyruvate kinase (Pyk), and transketolase (Tkt), were confirmed as surface expressed and all are able to interact with plasminogen. Plasminogen bound to recombinant proteins PdhB, GapA, and Pyk was converted to plasmin in the presence of urokinase plasminogen activator and plasmin-specific substrate d-valyl-leucyl-lysine-p-nitroanilide dihydrochloride. Furthermore, human fibrinogen was degraded by the complex of plasminogen and recombinant protein PdhB or Pgm. In addition, surface-displayed proteins (except PdhC) bind to human lung epithelial cells, and the interaction was reduced significantly by preincubation of cells with antiplasminogen. Our results suggest that plasminogen binding and activation by different surface-localized glycolytic enzymes of M. pneumoniae may play a role in successful and long-term colonization of the human respiratory tract.