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Analytical Biochemistry Jun 2024Prolidase (EC.3.4.13.9) is a dipeptidase known nowadays to play a pivotal role in several physiological and pathological processes. More in particular, this enzyme is... (Review)
Review
Prolidase (EC.3.4.13.9) is a dipeptidase known nowadays to play a pivotal role in several physiological and pathological processes. More in particular, this enzyme is involved in the cleavage of proline- and hydroxyproline-containing dipeptides (imidodipeptides), thus finely regulating the homeostasis of free proline and hydroxyproline. Abnormally high or low levels of prolidase have been found in numerous acute and chronic syndromes affecting humans (chronic liver fibrosis, viral and acute hepatitis, cancer, neurological disorders, inflammation, skin diseases, intellectual disability, respiratory infection, and others) for which the content of proline is well recognized as a clinical marker. As a consequence, the accurate analytical determination of prolidase activity is of greatly significant importance in clinical diagnosis and therapy. Apart from the Chinard's assay, some other more sensitive and well validated methodologies have been published. These include colorimetric and spectrophotometric determinations of free proline produced by enzymatic reactions, capillary electrophoresis, matrix-assisted laser desorption/ionization time-of-flight mass spectrometry, electrochemoluminescence, thin layer chromatography, and HPLC. The aim of this comprehensive review is to make a detailed survey of the in so far reported analytical techniques, highlighting their general features, as well as their advantages and possible drawbacks, providing in the meantime suggestions to stimulate further research in this intriguing field.
Topics: Humans; Colorimetry; Dipeptidases; Fibrosis; Hydroxyproline; Proline; Enzyme Assays
PubMed: 38460899
DOI: 10.1016/j.ab.2024.115506 -
Journal of the American Chemical Society Sep 2021Peptides constrained by intramolecular cross-links, especially stapled α-helices, have emerged as versatile scaffolds for drug development. However, there are fewer...
Peptides constrained by intramolecular cross-links, especially stapled α-helices, have emerged as versatile scaffolds for drug development. However, there are fewer examples of similarly constrained scaffolds for other secondary structures. Here, we used a novel computational strategy to identify an optimal staple for antiparallel β-strands, and then we incorporated that staple within a β-hairpin peptide. The hairpin uses 4-mercaptoproline as a novel staple component, which contributes to a unique, kinked structure. The stapled hairpins show a high degree of structure in aqueous solution, excellent resistance to degradation in cell lysates, and cytosolic penetration at micromolar concentrations. They also overlay with a unique subset of kinked hairpin motifs at protein-protein interaction interfaces. Thus, these scaffolds represent promising starting points for developing inhibitors of cellular protein-protein interactions.
Topics: Amino Acid Sequence; Models, Molecular; Peptides; Proline; Protein Structure, Secondary
PubMed: 34516087
DOI: 10.1021/jacs.1c04378 -
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 -
Journal of Computational Chemistry Jun 2021Computation of the thermodynamic consequences of protein mutations holds great promise in protein biophysics and design. Alchemical free energy methods can give improved...
Computation of the thermodynamic consequences of protein mutations holds great promise in protein biophysics and design. Alchemical free energy methods can give improved estimates of mutational free energies, and are already widely used in calculations of relative and absolute binding free energies in small molecule design problems. In principle, alchemical methods can address any amino acid mutation with an appropriate alchemical pathway, but identifying a strategy that produces such a path for proline and glycine mutations is an ongoing challenge. Most current strategies perturb only side chain atoms, while proline and glycine mutations also alter the backbone parameters and backbone ring topology. Some strategies also perturb backbone parameters and enable glycine mutations. This work presents a strategy that enables both proline and glycine mutations and comprises two key elements: a dual backbone with restraints and scaling of bonded terms, facilitating backbone parameter changes, and a soft bond in the proline ring, enabling ring topology changes in proline mutations. These elements also have utility for core hopping and macrocycle studies in computer-aided drug design. This new strategy shows slight improvements over an alternative side chain perturbation strategy for a set T4 lysozyme mutations lacking proline and glycine, and yields good agreement with experiment for a set of T4 lysozyme proline and glycine mutations not previously studied. To our knowledge this is the first report comparing alchemical predictions of proline mutations with experiment. With this strategy in hand, alchemical methods now have access to the full palette of amino acid mutations.
Topics: Glycine; Molecular Dynamics Simulation; Muramidase; Mutation; Proline; Thermodynamics
PubMed: 33844328
DOI: 10.1002/jcc.26525 -
Environmental Research May 2022Ozone is considered to be a major phytotoxic pollutant. It is an oxidizing molecule with harmful effects that can affect human health and vegetation. Due to its...
Ozone is considered to be a major phytotoxic pollutant. It is an oxidizing molecule with harmful effects that can affect human health and vegetation. Due to its phytotoxicity, it constitutes a threat to food security in a context of climate change. Proline accumulation is induced in response to numerous stresses and is assumed to be involved in plant antioxidant defense. We therefore addressed the question of the putative involvement of proline in plant ozone responses by analyzing the responses of two Arabidopsis mutants (obtained in the Col-0 genetic background) altered in proline metabolism and different ecotypes with various degrees of ozone sensitivity, to controlled ozone treatments. Among the mutants, the p5cs1 mutant plants accumulated less proline than the double prodh1xprodh2 (p1p2) mutants. Ozone treatments did not induce accumulation of proline in Col-0 nor in the mutant plants. However, the variation of the photosynthetic parameter Fv/Fm in the p1p2 mutant suggests a positive effect of proline. Proline accumulation induced by ozone was only observed in the most ozone-sensitive ecotypes, Cvi-0 and Ler. Contrary to our expectations, proline accumulation could not be correlated with variations in protein oxidation (carbonylation). On the other hand, flavonols content, measured here, using non-destructive methods, reflected exactly the genotypes ranking according to ozone sensitivity.
Topics: Arabidopsis Proteins; Flavonols; Gene Expression Regulation, Plant; Humans; Ozone; Proline
PubMed: 34662576
DOI: 10.1016/j.envres.2021.112214 -
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 -
Biochemical and Biophysical Research... Feb 2021Treatment of neurodegenerative diseases, such as Parkinson's disease, Huntington's chorea, Alzheimer's disease, is one of the priority directions in modern medicine....
BACKGROUND
Treatment of neurodegenerative diseases, such as Parkinson's disease, Huntington's chorea, Alzheimer's disease, is one of the priority directions in modern medicine. Thus, search and production of new physiologically active substances for the treatment of neurodegenerative disorders is one of the most important tasks for organic chemistry. The approach based on the replacement of a peptide bond in a peptide molecule with a structural isostere, non-hydrolyzable methylene phosphoryl fragment makes it possible to increase the metabolic stability of peptide molecules to the destructive action of peptidases.
METHODS
This work is devoted to the approbation of a new synthetic approach to the production of physiologically active substances in a series of peptide-type compounds with activity by replacing the peptide bond with isosteric methylene-phosphoryl fragment with the preservation of the original amino acid sequence.
RESULTS
A phosphine analog of the known physiologically active tripeptide proline-glycine-proline was obtained, cytotoxicity and neuroprotective properties of the initial tripeptide and its phosphine analog were studied.
CONCLUSION
Preliminary biological tests have shown that the obtained phosphine analog of the proline-glycine-proline tripeptide is involved in modulating the formation of sediments in the cellular system of proteinopathy, which may indicate their potential antiaggregatory properties.
Topics: Cell Line, Tumor; Humans; Neurodegenerative Diseases; Neuroprotective Agents; Oligopeptides; Phosphines; Proline; Protein Aggregation, Pathological
PubMed: 33412416
DOI: 10.1016/j.bbrc.2020.12.087 -
The Journal of Organic Chemistry May 2019Organocatalysis is an emerging field, in which small metal-free organic structures catalyze a diversity of reactions with a remarkable stereoselectivity. The ability to...
Organocatalysis is an emerging field, in which small metal-free organic structures catalyze a diversity of reactions with a remarkable stereoselectivity. The ability to selectively switch on such pathways upon demand has proven to be a valuable tool in biological systems. Light as a trigger provides the ultimate spatial and temporal control of activation. However, there have been limited examples of phototriggered catalytic systems. Herein, we describe the synthesis and application of a caged proline system that can initiate organocatalysis upon irradiation. The caged proline was generated using the highly efficient 4-carboxy-5,7-dinitroindolinyl (CDNI) photocleavable protecting group in a four-step synthesis. Advantages of this system include water solubility, biocompatibility, high quantum yield for catalyst release, and responsiveness to two-photon excitation. We showed the light-triggered catalysis of a crossed aldol reaction, a Mannich reaction, and a self-aldol condensation reaction. We also demonstrated light-initiated catalysis, leading to the formation of a biocide in situ, which resulted in the growth inhibition of E. coli, with as little as 3 min of irradiation. This technique can be broadly applied to other systems, by which the formation of active forms of drugs can be catalytically assembled remotely via two-photon irradiation.
Topics: Anti-Bacterial Agents; Catalysis; Escherichia coli; Indoles; Kinetics; Photochemical Processes; Proline; Solubility; Water
PubMed: 30908906
DOI: 10.1021/acs.joc.9b00220 -
The Journal of Organic Chemistry Mar 2019Fluorinated proline derivatives have found diverse applications in areas ranging from medicinal chemistry over structural biochemistry to organocatalysis. Depending on...
Fluorinated proline derivatives have found diverse applications in areas ranging from medicinal chemistry over structural biochemistry to organocatalysis. Depending on the stereochemistry of monofluorination at the proline 3- or 4-position, different effects on the conformational properties of proline (ring pucker, cis/ trans isomerization) are introduced. With fluorination at both 3- and 4-positions, matching or mismatching effects can occur depending on the relative stereochemistry. Here we report, in full, the syntheses and conformational properties of three out of the four possible 3,4-difluoro-l-proline diastereoisomers. The yet unreported conformational properties are described for (3 S,4 S)- and (3 R,4 R)-difluoro-l-proline, which are shown to bias ring pucker and cis/ trans ratios on the same order of magnitude as their respective monofluorinated progenitors, although with significantly faster amide cis/ trans isomerization rates. The reported analogues thus expand the scope of available fluorinated proline analogues as tools to tailor proline's distinct conformational and dynamical properties, allowing for the interrogation of its role in, for instance, protein stability or folding.
Topics: Halogenation; Molecular Conformation; Proline; Stereoisomerism
PubMed: 30777755
DOI: 10.1021/acs.joc.8b02920 -
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