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Biochemistry Nov 2021Thiazolidine carboxylates such as thiazolidine-4-carboxylate (T4C) and thiazolidine-2-carboxylate (T2C) are naturally occurring sulfur analogues of proline. These...
Thiazolidine carboxylates such as thiazolidine-4-carboxylate (T4C) and thiazolidine-2-carboxylate (T2C) are naturally occurring sulfur analogues of proline. These compounds have been observed to have both beneficial and toxic effects in cells. Given that proline dehydrogenase has been proposed to be a key enzyme in the oxidative metabolism of thioprolines, we characterized T4C and T2C as substrates of proline catabolic enzymes using proline utilization A (PutA), which is a bifunctional enzyme with proline dehydrogenase (PRODH) and l-glutamate-γ-semialdehyde dehydrogenase (GSALDH) activities. PutA is shown here to catalyze the FAD-dependent PRODH oxidation of both T4C and T2C with catalytic efficiencies significantly higher than with proline. Stopped-flow experiments also demonstrate that l-T4C and l-T2C reduce PutA-bound FAD at rates faster than proline. Unlike proline, however, oxidation of T4C and T2C does not generate a substrate for NAD-dependent GSALDH. Instead, PutA/PRODH oxidation of T4C leads to cysteine formation, whereas oxidation of T2C generates an apparently stable Δ-thiazoline-2-carboxylate species. Our results provide new insights into the metabolism of T2C and T4C.
Topics: Bacterial Proteins; Cysteine; Enzyme Assays; Kinetics; Membrane Proteins; Proline; Recombinant Proteins; Sinorhizobium meliloti; Thiazolidines
PubMed: 34752700
DOI: 10.1021/acs.biochem.1c00625 -
Molecules (Basel, Switzerland) Dec 2021The 3D structure and surface characteristics of proteins and peptides are crucial for interactions with receptors or ligands and can be modified to some extent to... (Review)
Review
The 3D structure and surface characteristics of proteins and peptides are crucial for interactions with receptors or ligands and can be modified to some extent to modulate their biological roles and pharmacological activities. The introduction of halogen atoms on the side-chains of amino acids is a powerful tool for effecting this type of tuning, influencing both the physico-chemical and structural properties of the modified polypeptides, helping to first dissect and then rationally modify features that affect their mode of action. This review provides examples of the influence of different types of halogenation in amino acids that replace native residues in proteins and peptides. Examples of synthetic strategies for obtaining halogenated amino acids are also provided, focusing on some representative compounds and their biological effects. The role of halogenation in native and designed antimicrobial peptides (AMPs) and their mimetics is then discussed. These are in the spotlight for the development of new antimicrobial drugs to counter the rise of antibiotic-resistant pathogens. AMPs represent an interesting model to study the role that natural halogenation has on their mode of action and also to understand how artificially halogenated residues can be used to rationally modify and optimize AMPs for pharmaceutical purposes.
Topics: Anti-Bacterial Agents; Antimicrobial Peptides; Gram-Negative Bacteria; Gram-Positive Bacteria; Halogenation; Halogens; Humans; Microbial Sensitivity Tests; Peptidomimetics; Peptoids; Proline; Structure-Activity Relationship
PubMed: 34885985
DOI: 10.3390/molecules26237401 -
Scientific Reports Nov 2022Overcoming the skin barrier properties efficiently, temporarily, and safely for successful transdermal drug delivery remains a challenge. We synthesized three series of...
Overcoming the skin barrier properties efficiently, temporarily, and safely for successful transdermal drug delivery remains a challenge. We synthesized three series of potential skin permeation enhancers derived from natural amino acid derivatives proline, 4-hydroxyproline, and pyrrolidone carboxylic acid, which is a component of natural moisturizing factor. Permeation studies using in vitro human skin identified dodecyl prolinates with N-acetyl, propionyl, and butyryl chains (Pro2, Pro3, and Pro4, respectively) as potent enhancers for model drugs theophylline and diclofenac. The proline derivatives were generally more active than 4-hydroxyprolines and pyrrolidone carboxylic acid derivatives. Pro2-4 had acceptable in vitro toxicities on 3T3 fibroblast and HaCaT cell lines with IC values in tens of µM. Infrared spectroscopy using the human stratum corneum revealed that these enhancers preferentially interacted with the skin barrier lipids and decreased the overall chain order without causing lipid extraction, while their effects on the stratum corneum protein structures were negligible. The impacts of Pro3 and Pro4 on an in vitro transepidermal water loss and skin electrical impedance were fully reversible. Thus, proline derivatives Pro3 and Pro4 have an advantageous combination of high enhancing potency, low cellular toxicity, and reversible action, which is important for their potential in vivo use as the skin barrier would quickly recover after the drug/enhancer administration is terminated.
Topics: Humans; Skin Absorption; Hydroxyproline; Proline; Permeability; Administration, Cutaneous; Skin; Pharmaceutical Preparations; Organic Chemicals; Pyrrolidinones; Carboxylic Acids
PubMed: 36376455
DOI: 10.1038/s41598-022-24108-6 -
Anaerobe Jun 2020Clostridioides difficile colonizes the intestines of susceptible individuals and releases toxins that mediate disease. To replicate and expand in the intestines,...
Clostridioides difficile colonizes the intestines of susceptible individuals and releases toxins that mediate disease. To replicate and expand in the intestines, C. difficile ferments proline, and this activity is influenced by the availability of proline and trace nutrients. C. difficile must also compete with the commensal microbiota for these limited nutrients. The specific microbes present in the intestines that may shape the ability of C. difficile to benefit from proline fermentation are unknown. In this study we developed a panel of commensal Clostridia to test the hypothesis that the microbiota influences C. difficile growth through proline fermentation. The experimental panel of Clostridia was composed of murine and human isolates that ranged in their capacity to ferment proline in different media. Competition between wild type C. difficile and a mutant strain unable to ferment proline (prdB:CT) in the presence of these Clostridia revealed that bacteria closely related to Paraclostridium benzoelyticum and Paeniclostridium spp. decreased the benefit to C. difficile provided by proline fermentation. Conversely, Clostridium xylanolyticum drove C. difficile towards an increased reliance on proline fermentation for growth. Overall, the ability of C. difficile to benefit from proline fermentation is contextual and in part dependent on the microbiota.
Topics: Animals; Antibiosis; Clostridiaceae; Clostridiales; Gastrointestinal Microbiome; Humans; Mice; Proline
PubMed: 32422411
DOI: 10.1016/j.anaerobe.2020.102210 -
The FEBS Journal Jul 2021Linker for activation in T cells (LAT) is a critical regulator of T-cell development and function. It organises signalling events at the plasma membrane. However, the...
Linker for activation in T cells (LAT) is a critical regulator of T-cell development and function. It organises signalling events at the plasma membrane. However, the mechanism, which controls LAT localisation at the plasma membrane, is not fully understood. Here, we studied the impact of helix-breaking amino acids, two prolines and one glycine, in the transmembrane segment on localisation and function of LAT. Using in silico analysis, confocal and super-resolution imaging and flow cytometry, we demonstrate that central proline residue destabilises transmembrane helix by inducing a kink. The helical structure and dynamics are further regulated by glycine and another proline residue in the luminal part of LAT transmembrane domain. Replacement of these residues with aliphatic amino acids reduces LAT dependence on palmitoylation for sorting to the plasma membrane. However, surface expression of these mutants is not sufficient to recover function of nonpalmitoylated LAT in stimulated T cells. These data indicate that geometry and dynamics of LAT transmembrane segment regulate its localisation and function in immune cells.
Topics: Adaptor Proteins, Signal Transducing; Amino Acid Sequence; Calcium; Cell Membrane; Glycine; Humans; Jurkat Cells; Membrane Proteins; Microscopy, Confocal; Microscopy, Interference; Molecular Dynamics Simulation; Mutation; Proline; Protein Domains; Protein Structure, Secondary; Sequence Homology, Amino Acid; T-Lymphocytes
PubMed: 33458942
DOI: 10.1111/febs.15713 -
Microbial Cell Factories Aug 2022In recent years, there has been a growing demand for microbial production of trans-4-hydroxy-L-proline (t4Hyp), which is a value-added amino acid and has been widely...
BACKGROUND
In recent years, there has been a growing demand for microbial production of trans-4-hydroxy-L-proline (t4Hyp), which is a value-added amino acid and has been widely used in the fields of medicine, food, and cosmetics. In this study, a multivariate modular metabolic engineering approach was used to remove the bottleneck in the synthesis pathway of t4Hyp.
RESULTS
Escherichia coli t4Hyp synthesis was performed using two modules: a α-ketoglutarate (α-KG) synthesis module (K module) and L-proline synthesis with hydroxylation module (H module). First, α-KG attrition was reduced, and then, L-proline consumption was inhibited. Subsequently, to improve the contribution to proline synthesis with hydroxylation, optimization of gene overexpression, promotor, copy number, and the fusion system was performed. Finally, optimization of the H and K modules was performed in combination to balance metabolic flow. Using the final module H1K4 in a shaking flask culture, 8.80 g/L t4Hyp was produced, which was threefold higher than that produced by the W0 strain.
CONCLUSIONS
These strategies demonstrate that a microbial cell factory can be systematically optimized by modular engineering for efficient production of t4Hyp.
Topics: Bacterial Outer Membrane Proteins; Escherichia coli; Escherichia coli Proteins; Hydroxyproline; Ketoglutaric Acids; Metabolic Engineering; Proline
PubMed: 35953819
DOI: 10.1186/s12934-022-01884-4 -
Communications Biology Sep 2022Hyperprolinemia type II (HPII) is an inborn error of metabolism due to genetic variants in ALDH4A1, leading to a deficiency in Δ-1-pyrroline-5-carboxylate (P5C)...
Hyperprolinemia type II (HPII) is an inborn error of metabolism due to genetic variants in ALDH4A1, leading to a deficiency in Δ-1-pyrroline-5-carboxylate (P5C) dehydrogenase. This leads to an accumulation of toxic levels of P5C, an intermediate in proline catabolism. The accumulating P5C spontaneously reacts with, and inactivates, pyridoxal 5'-phosphate, a crucial cofactor for many enzymatic processes, which is thought to be the pathophysiological mechanism for HPII. Here, we describe the use of a combination of LC-QTOF untargeted metabolomics, NMR spectroscopy and infrared ion spectroscopy (IRIS) to identify and characterize biomarkers for HPII that result of the spontaneous reaction of P5C with malonic acid and acetoacetic acid. We show that these biomarkers can differentiate between HPI, caused by a deficiency of proline oxidase activity, and HPII. The elucidation of their molecular structures yields insights into the disease pathophysiology of HPII.
Topics: 1-Pyrroline-5-Carboxylate Dehydrogenase; Amino Acid Metabolism, Inborn Errors; Biomarkers; Phosphates; Proline; Proline Oxidase; Pyridoxal; Pyrroles
PubMed: 36131087
DOI: 10.1038/s42003-022-03960-2 -
Scientific Reports May 2023This work was conducted to study positive and negative impacts of cerium (Ce) and samarium (Sm) on two cultivars (Arta and Baharan) in wheat plant. Symbols of stress...
This work was conducted to study positive and negative impacts of cerium (Ce) and samarium (Sm) on two cultivars (Arta and Baharan) in wheat plant. Symbols of stress such as proline, malondialdehyde (MDA) and antioxidant enzymes, which may be complicated in the suppression responses of plants, were also studied. Wheat plants were exposed to 0, 2500, 5000, 7500, 10,000 and 15,000 μM of Ce and Sm for 7 days. The growth enhanced in plants treated with lesser Ce and Sm concentration (2500 μM) and declined in plants treated with upper concentrations as compared to untreated plants. The treatment with 2500 μM of Ce and Sm increased dry weigh in Arta by 68.42 and 20% and in Baharan by 32.14% and 27.3%. Thus, Ce and Sm had hormesis effect on growth in wheat plants. According to plant's growth parameter patterns, Arta cultivar had more sensitive to Sm than to Ce, whereas Baharan cultivar had sensitive to Ce than to Sm. Our results indicated impact of Ce and Sm on proline accumulation depended on the dosage of Ce and Sm. It was observed that Ce and Sm accumulated in wheat plants at higher exposure doses. Increment of MDA content by Ce and Sm treatments showed that these metals caused oxidative stress in wheat plants. Ce and Sm blocked enzymatic antioxidant system (superoxide dismutases, peroxidase and polyphenol peroxidase) in wheat. In wheat plants treated with lower Ce and Sm concentrations higher amounts of non-enzymatic antioxidant metabolites were detected. Thus, we showed the potential negative impact of unsuitable utilization of REEs in plants and suggested growth and interruption in physiological and biochemical mechanisms as a possible factor to recognize the underlying toxicological processes.
Topics: Antioxidants; Triticum; Samarium; Cerium; Peroxidase; Plants; Proline
PubMed: 37217773
DOI: 10.1038/s41598-023-35561-2 -
Journal of the American Society For... Aug 2023The fragmentation characteristics of ions produced from proline-containing heptapeptides have been studied in detail. The study has utilized the following C-terminally...
The fragmentation characteristics of ions produced from proline-containing heptapeptides have been studied in detail. The study has utilized the following C-terminally amidated model peptides: PA, APA, APA, APA, APA, APA, AP, PYAGFLV, PAGFLVY, PGFLVYA, PFLVYAG, PLVYAGF, PVYAGFL, YPAGFLV, YAPGFLV, YAGPFLV, YAGFPLV, YAGFLPV, YAGFLVP, PYAFLVG, PVLFYAG, APXA, and AXPA (where X = C, D, F, G, L, V, and Y, respectively). The results have shown that ions undergo head-to-tail cyclization and form a macrocyclic structure. Under the collision-induced dissociation (CID) condition, it generates nondirect sequence ions regardless of the position of the proline and the neighboring amino acid residues. This study highlights the unusual and unique fragmentation behavior of proline-containing heptapeptides. Following the head-to-tail cyclization, the ring opens up and places the proline residue in the N-terminal position while forming a regular oxazolone form of ions for all peptide series. Then, the fragmentation reaction pathway is followed by the elimination of proline with its C-terminal neighbor residue as an oxazolone (e.g., PX) for all proline-containing peptide series.
Topics: Oxazolone; Proline; Peptides; Ions; Cyclization
PubMed: 37402129
DOI: 10.1021/jasms.3c00049 -
Lancet (London, England) Feb 2022
Topics: COVID-19 Testing; Developing Countries; Drug Combinations; Global Health; Health Services Accessibility; Humans; Lactams; Leucine; Nitriles; Proline; Ritonavir; COVID-19 Drug Treatment
PubMed: 35219388
DOI: 10.1016/S0140-6736(22)00372-5