Oral glutamine dipeptide or oral glutamine free amino acid reduces burned injury progression in rats

This study was carried out to evaluate the effect of Glutamine, as a dipeptide or a free amino acid form, on the progression of burn injuries in rats. Thirty male Wistar rats were burned with a comb metal plate heated in boiling water (98 °C) for three minutes, creating four rectangular full-thickness burn areas separated by three unburned interspaces (zone of stasis) in both dorsum sides. The animals were randomized into three groups (n=10): saline solution (G1-Control) and treated groups that orally received Glutamine as dipeptide (G2-Dip) or free amino acid (G3-FreeAA). Two and seven days after burn injury, lesions were photographed for unburned interspaces necrosis evolution assessment. Seven days after injury, glutathione seric was measured and histopathological analysis was performed. By photographs, there was a significant reduction in necrosis progression in G3-Free-AA between days two and seven. Histopathological analysis at day 7 showed a significantly higher stasis zone without necrosis and a higher number of fibroblasts in G2-Dip and G3-FreeAA compared with G1-Control. Also, glutathione serum dosage was higher in G2-Dip. The plasmatic glutathione levels were higher in the G2-Dip than the G1-Control, and there was a trend to higher levels in G3-FreeAA. The reduction in histological lesions, greater production of fibroblasts, and greater amounts of glutathione may have benefited the evolution of burn necrosis, which showed greater preservation of interspaces.

The Amyloid Fibril-Forming β-Sheet Regions of Amyloid β and α-Synuclein Preferentially Interact with the Molecular Chaperone 14-3-3ζ

14-3-3 proteins are abundant, intramolecular proteins that play a pivotal role in cellular signal transduction by interacting with phosphorylated ligands. In addition, they are molecular chaperones that prevent protein unfolding and aggregation under cellular stress conditions in a similar manner to the unrelated small heat-shock proteins. In vivo, amyloid β (Aβ) and α-synuclein (α-syn) form amyloid fibrils in Alzheimer’s and Parkinson’s diseases, respectively, a process that is intimately linked to the diseases’ progression. The 14-3-3ζ isoform potently inhibited in vitro fibril formation of the 40-amino acid form of Aβ (Aβ40) but had little effect on α-syn aggregation. Solution-phase NMR spectroscopy of 15N-labeled Aβ40 and A53T α-syn determined that unlabeled 14-3-3ζ interacted preferentially with hydrophobic regions of Aβ40 (L11-H21 and G29-V40) and α-syn (V3-K10 and V40-K60). In both proteins, these regions adopt β-strands within the core of the amyloid fibrils prepared in vitro as well as those isolated from the inclusions of diseased individuals. The interaction with 14-3-3ζ is transient and occurs at the early stages of the fibrillar aggregation pathway to maintain the native, monomeric, and unfolded structure of Aβ40 and α-syn. The N-terminal regions of α-syn interacting with 14-3-3ζ correspond with those that interact with other molecular chaperones as monitored by in-cell NMR spectroscopy.

Rules for designing protein fold switches and their implications for the folding code

We have engineered switches between the three most common small folds, 3a, 4b+a, and a/b plait, referred to here as A, B, and S, respectively. Mutations were introduced into the natural S protein until sequences were created that have a stable S-fold in their longer (~90 amino acid) form and have an alternative fold (either A or B) in their shorter (56 amino acid) form. Five sequence pairs were designed and key structures were determined using NMR spectroscopy. Each protein pair is 100% identical in the 56 amino acid region of overlap. Several rules for engineering switches emerged. First, designing one sequence with good native state interactions in two folds requires care but is feasible. Once this condition is met, fold populations are determined by the stability of the embedded A- or B-fold relative to the S-fold and the conformational propensities of the ends that are generated in the switch to the embedded fold. If the stabilities of the embedded fold and the longer fold are similar, conformation is highly sensitive to mutation so that even a single amino acid substitution can radically shift the population to the alternative fold. The results provide insight into why dimorphic sequences can be engineered and sometimes exist in nature, while most natural protein sequences populate single folds. Proteins may evolve toward unique folds because dimorphic sequences generate interactions that destabilize and can produce aberrant functions. Thus two-state behavior may result from nature's negative design rather than being an inherent property of the folding code.

The evolution of Genetic Molecular Map and phylogenetic tree of Coronavirus (COVID-19)

Coronavirus (COVID-19) is suspected to originate from an animal host (zoonotic) followed by a human to human transmission, The purpose of the present study is to determine the genetic affinity of a phylogenetic tree and conformation of protein between human and bat. Through study of genetic sequencing, as shown in the tree design of strains and genetic variants, the main cause of COVID-19 is the Bat Coronavirus RaTG13 (SARS) virus in ID: MN996532.1 and ID: MG772933.1 (24-Jul-2013 and Feb-2017) shows that the evolution of the Corona virus from 2003, 2013 to 2020 which has become the most deadly peak in humans, the virus evolved from the bat effect on Humans. Protein analysis show 98 change of amino acid form RaTG13 (SARS) virus to COVID-19 Homo sapiens. Consequently, this study increased our understanding of the genetic variety of the COVID-19 carried by bats. So we conclude that a protein conformation drawing shows high identity compatibility between a bat and a human

Hydrophobic Amino Acid Tryptophan Shows Promise as a Potential Absorption Enhancer for Oral Delivery of Biopharmaceuticals

Cell-penetrating peptides (CPPs) have great potential to efficiently deliver drug cargos across cell membranes without cytotoxicity. Cationic arginine and hydrophobic tryptophan have been reported to be key component amino acids for cellular internalization of CPPs. We recently found that l-arginine could increase the oral delivery of insulin in its single amino acid form. Therefore, in the present study, we evaluated the ability of another key amino acid, tryptophan, to enhance the intestinal absorption of biopharmaceuticals. We demonstrated that co-administration with l-tryptophan significantly facilitated the oral and intestinal absorption of the peptide drug insulin administered to rats. Furthermore, l-tryptophan exhibited the ability to greatly enhance the intestinal absorption of other peptide drugs such as glucagon-like peptide-1 (GLP-1), its analog Exendin-4 and macromolecular hydrophilic dextrans with molecular weights ranging from 4000 to 70,000 g/mol. However, no intermolecular interaction between insulin and l-tryptophan was observed and no toxic alterations to epithelial cellular integrity—such as changes to cell membranes, cell viability, or paracellular tight junctions—were found. This suggests that yet to be discovered inherent biological mechanisms are involved in the stimulation of insulin absorption by co-administration with l-tryptophan. These results are the first to demonstrate the significant potential of using the single amino acid l-tryptophan as an effective and versatile bioavailability enhancer for the oral delivery of biopharmaceuticals.

Differential processing of pro-glucose-dependent insulinotropic polypeptide in gut

Glucose-dependent insulinotropic polypeptide (GIP) is a hormone released from enteroendocrine K cells in response to meals. Posttranslational processing of the precursor protein pro-GIP at residue 65 by proprotein convertase subtilisin/kexin type 1 (PC1/3) in gut K cells gives rise to the established 42-amino-acid form of GIP (GIP1–42). However, the pro-GIP peptide sequence contains a consensus cleavage site for PC2 at residues 52–55 and we identified PC2 immunoreactivity in a subset of K cells, suggesting the potential existence of a COOH-terminal truncated GIP isoform, GIP1–30. Indeed a subset of mouse and human K cells display GIP immunoreactivity with GIP antibodies directed to the mid portion of the peptide, but not with a COOH-terminal-directed GIP antibody, indicative of the presence of a truncated form of GIP. This population of cells represents ∼5–15% of the total GIP-immunoreactive cells in mice, depending on the region of intestine, and is virtually absent in mice lacking PC2. Amidated GIP1–30 and GIP1–42 have comparable potency at stimulating somatostatin release in the perfused mouse stomach. Therefore, GIP1–30 represents a naturally occurring, biologically active form of GIP.

Confounding Factors Influencing Amyloid Beta Concentration in Cerebrospinal Fluid

Background. Patients afflicted with Alzheimer's disease (AD) exhibit a decrease in the cerebrospinal fluid (CSF) concentration of the 42 amino acid form ofβ-amyloid (Aβ42). However, a high discrepancy between different centers in measuredAβ42levels reduces the utility of this biomarker as a diagnostic tool and in monitoring the effect of disease modifying drugs. Preanalytical and analytical confounding factors were examined with respect to their effect on the measuredAβ42level.Methods. Aliquots of CSF samples were either treated differently prior toAβ42measurement or analyzed using different commercially available xMAP or ELISA assays.Results. Confounding factors affecting CSFAβ42levels were storage in different types of test tubes, dilution with detergent-containing buffer, plasma contamination, heat treatment, and the origin of the immunoassays used for quantification.Conclusion. In order to conduct multicenter studies, a standardized protocol to minimize preanalytical and analytical confounding factors is warranted.