Acid Position Research Articles

Electrostatic Preorganization in Three Distinct Heterogeneous Proteasome β-Subunits.

The origin of the enzyme's powerful role in accelerating chemical reactions is one of the most critical and still widely discussed questions. It is already accepted that enzymes impose an electrostatic field onto their substrates by adopting complex three-dimensional structures; therefore, the preorganization of electric fields inside protein active sites has been proposed as a crucial contributor to catalytic mechanisms and rate constant enhancement. In this work, we focus on three catalytically active β-subunits of 20S proteasomes with low sequence identity (∼30%) whose active sites, although situated in an electrostatically miscellaneous environment, catalyze the same chemical reaction with similar catalytic efficiency. Our in silico experiments reproduce the experimentally observed equivalent reactivity of the three sites and show that obliteration of the electrostatic potential in all active sites would deprive the enzymes of their catalytic power by slowing down the chemical process by a factor of 1035. To regain enzymatic efficiency, besides catalytic Thr1 and Lys33 residues, the presence of aspartic acid in position 17 and an aqueous solvent is required, proving that the electrostatic potential generated by the remaining residues is insignificant for catalysis. Moreover, it was found that the gradual decay of atomic charges on Asp17 strongly correlates with the enzyme's catalytic rate deterioration as well as with a change in the charge distributions due to introduced mutations. The computational procedure used and described here may help identify key residues for catalysis in other biomolecular systems and consequently may contribute to the process of designing enzyme-like synthetic catalysts.

Three-Dimensional Determination (Morphometric Study) of Arch Perimeter and Palatal Depth in an Iraqi Sample Aged 17–25 Years with Sickle Cell Anemia: A Cross-Sectional Study

Sickle cell anemia (SCA) is a hereditary disorder caused by the substitution of valine for glutamic acid in position six of globin's beta chain. Segmental arch dimensions, arch perimeter, and palatal depth comparison between SCA and control using the three-dimensional (3D) technique was the aim of this study. This cross-sectional investigation was carried out in Karbala City. The sample consisted of 200 casts (100 SCA and 100 control). Each group consisted of 50 casts upper, and 50 casts lower (25 males and 25 females). The arch perimeter was calculated using two segmented arch lengths bilaterally. Palatal depth was measured from the mesiolingual cusp tip of the upper first permanent molar to the corresponding palatal vault. The difference between the control and SCA groups was calculated using the independent t-test. Except for lower left incisal to canine distance (LLI-C), where p = 0.01 there was no significant difference in segmental arch measurements and perimeter between males in SCA and males in control. There was no significant difference in segmental arch measurements and perimeter between females in SCA and females in control. There was no significant difference in palatal depth between the control and SCA (male and female). SCA did not affect the segmental, arch perimeter, and palatal depth measurements.

Synthesis and Biological Studies of New Temporin A Analogs Containing Unnatural Amino Acids in Position 7.

(1) Background: Antimicrobial resistance is growing at an extreme pace and has proven to be an urgent topic, for research into alternative treatments. Such a prospective possibility is hidden in antimicrobial peptides because of their low to no toxicity, effectiveness at low concentrations, and most importantly their ability to be used for multiple treatments. This work was focused on the study of the effect of the modification in position 7 of Temporin A on its biological activity; (2) Methods: The targeted peptides were synthesized using Fmoc/Ot-Bu SPPS. The antibacterial activity of the analogs was determined using the broth microdilution method and disk-diffusion method. In vitro tests were performed to determine the cytotoxicity, phototoxicity, and antiproliferative activity of the peptide analogs on a panel of tumor and normal cell lines; (3) Results: All analogs except DTCit showed good antibacterial activity, with DTDab having the best activity according to the disk-diffusion method. However, DTCit had an acceptable cytotoxicity, combined with good selectivity against the test MCF-7 cell line; (4) Conclusions: The obtained results revealed the importance of the basicity and length of the side chain at position 7 in the Temporin A sequence for both tested activities.

Selectivity Switch in Non‐Directed Palladium‐Catalyzed C−H Olefination of Chlorobenzene Derivatives

AbstractA palladium(II)‐catalyzed oxidative C−H olefination of chlorobenzene derivatives, which favored the functionalization at the ortho‐to‐chlorine position, was reported. The catalytic system employed a commercially available quinoline ligand to access a wide panel of olefinated aryl chlorides in moderate to high yields from feedstock chemicals (42 examples, 28 to 93% yields). The synthetic utility of the method was further demonstrated by the coupling of various bio‐relevant acrylate partners offering straightforward access to structurally diverse complex molecules. A computational study of the reaction allowed us to confirm that the C−H activation step, occurring through a Concerted Metalation‐Deprotonation mechanism, determined the site‐selectivity of the transformation. The regioselectivity was controlled by the pKa of the C−H bond (more acidic position) and the lower distortion of chlorobenzene in the key transition structure.

Unusual 1-3 peptidoglycan cross-links in Acetobacteraceae are made by L,D-transpeptidases with a catalytic domain distantly related to YkuD domains

Peptidoglycan is an essential component of the bacterial cell envelope that contains glycan chains substituted by short peptide stems. Peptide stems are polymerized by D,D-transpeptidases, which make bonds between the amino acid in position four of a donor stem and the third residue of an acceptor stem (4-3 cross-links). Some bacterial peptidoglycans also contain 3-3 cross-links that are formed by another class of enzymes called L,D-transpeptidases which contain a YkuD catalytic domain. In this work, we investigate the formation of unusual bacterial 1-3 peptidoglycan cross-links. We describe a version of the PGFinder software that can identify 1-3 cross-links and report the high-resolution peptidoglycan structure of Gluconobacter oxydans (a model organism within the Acetobacteraceae family). We reveal that G.oxydans peptidoglycan contains peptide stems made of a single alanine as well as several dipeptide stems with unusual amino acids at their C-terminus. Using a bioinformatics approach, we identified aG.oxydans mutant from a transposon library with a drastic reduction in 1-3 cross-links. Through complementation experiments in G.oxydans and recombinant protein production in a heterologous host, we identify an L,D-transpeptidase enzyme with a domain distantly related to the YkuD domain responsible for these non-canonical reactions. This work revisits the enzymatic capabilities of L,D-transpeptidases, a versatile family of enzymes that play a key role in bacterial peptidoglycan remodelling.

Genomic Basis of Adaptive Divergence in Leg Length between Ground- and Tree-Dwelling Species within a Bird Family.

Hind limbs of tetrapods vary greatly in length and the variability can be associated with locomotor adaptation. Although the phenotypic evolution has been well documented, the underlying genetic basis remains poorly understood. We address this issue by integrating comparative genomics and functional prediction with a study system consisting of ground-dwelling, long-legged and tree-dwelling, short-legged species within the avian family Paridae. Genome-wide divergence and phenotypic correlation analyses jointly identified five highly divergent genomic regions that are significantly related with the difference in leg length between these two groups. Gene annotation for these regions detected three genes involved in skeletal development, that is, PTPA, BRINP1, and MIGA2, with the first one being under the strongest selection. Furthermore, four single nucleotide polymorphisms (SNPs) in the coding region of PTPA can well distinguish the two groups with distinct leg length. Among the four SNPs, one is non-synonymous mutation, and according to the prediction for protein structure and function, it can affect the 3D structure of the encoded protein by altering the corresponding amino acid's position. The alleles of PTPA were found in all sequenced species of the orders Palaeognathae and Psittaciformes, which typically take a ground locomotion style. A whole-genome scanning across bird species uncovered that the four SNPs are more likely to be present in resident passerines with increased leg length/wing length ratios (a proxy of leg-dependent locomotion efficiency). Our findings provide insight into the molecular evolution of locomotion performance based on leg morphology in birds.

Two granzyme A/K homologs in Zebra mbuna have different specificities, one classical tryptase and one with chymase activity

Granzymes A and K are two highly homologous serine proteases expressed by mammalian cytotoxic T cells (CTLs) and natural killer (NK) cells. The locus encoding these two proteases is the first of the hematopoietic serine protease loci to appear during vertebrate evolution. This locus is found in all jawed vertebrates including the cartilaginous fishes. Granzyme A is the most abundant of the different granzymes expressed by CTLs and NK cells and its potential function has been studied extensively for many years. However, no clear conclusions concerning its primary role in the immune defense has been obtained. In all mammals, there are only one copy each of granzyme A and K, whereas additional copies are found in both cartilaginous and ray finned fishes. In cichlids two of these copies seem to encode new members of the granzyme A/K family. These two new members appear to have changed primary specificity and to be pure chymases based on the amino acids in their active site substrate binding pockets. Interestingly, one of these gene copies is located in the middle of the granzyme A/K locus, while the other copy is present in another locus, the met-ase locus. We here present a detailed characterization of the extended cleavage specificity of one of these non-classical granzymes, a Zebra mbuna granzyme positioned in the granzyme A/K locus. This enzyme, named granzyme A2, showed a high preference for tyrosine in the P1 position of substrates, thereby being a strict chymase. We have also characterized one of the classical granzyme A/Ks of the Zebra mbuna, granzyme A1, which is a tryptase with preference for arginine in the P1 position of substrates. Based on their extended specificities, the two granzymes showed major similarities, but also some differences in preferred amino acids in positions surrounding the cleavable amino acid. Fish lack one of the hematopoietic serine protease loci of mammals, the chymase locus, where one of the major mast cell enzymes is located. An interesting question is now if cichlids have by compensatory mechanisms generated a mast cell chymase from another locus, and if similar chymotryptic enzymes have appeared also in other fish species.

The Positional Distribution of Fatty Acids in Palm Oil and Lard Influences Their Biologic Effects in Rats

Dietary fatty acids in the sn-2 position are preferentially absorbed as monoacylglycerols. To determine whether they also have more important biologic effects, rats were fed for 2 and 4 mo a purified diet containing native palm oil, interesterified palm oil, native lard or interesterified lard. Interesterification that increased or decreased the level of fatty acids in position 2 depending on the fat, resulted in significant corresponding changes in the fecal excretion of saturated fatty acids. Fecal excretion of saturated fatty acids was associated with significant changes in some plasma fatty acids. Interesterification in lard resulted in significantly lower plasma triglycerides, and in palm oil, increased platelet aggregation induced by ADP. Lipemia, platelet aggregation and associated plasma fatty acids (palmitic, heneicosanoic and docosahexaenoic acids) were significantly affected only by dietary fatty acids at the sn-2 position. Even without changes in absorption, only linoleic acid in position 2 was correlated with the plasma concentration of the corresponding longer chain arachidonic acid. These results in rats confirm that the fatty acids (saturated and unsaturated) in position 2 of dietary triglycerides play a crucial role in the metabolism and biologic effects of these fatty acids.

Detection of the SARS-CoV-2 Omicron Variant in COVID-19 Patients from South Tangerang Using SNP-Probes S371L and K417N

The COVID-19 pandemic caused by the SARS-CoV-2 virus has posed a global challenge. Experts from various branches of science have endeavoured to find solutions to control its spread, one of which has been the quick and precise detection of the virus and its variants in patients. This study aimed to detect the presence of SARS-CoV-2, notably the rapidly spreading Omicron variant, using the spike (S)-gene target failure (SGTF) and S-gene target positive (SGTP) with the principle of the single nucleotide polymorphism (SNP)-probe test. Our descriptive experimental approach detected Omicron variants with the SNP-probe technique using samples of SARS-CoV-2 patients and controls. The probes were designed to recognize the nucleotide code of the amino acids in positions 371 and 417 of SARS-CoV-2. The existence of variants was monitored by the presence or absence of a fluorescence signal, which was translated into a sigmoidal graph using a real-time (RT)-PCR machine. One hundred and twelve samples that had tested positive for SARS-CoV-2 and the Omicron variant using a registered commercial kit showed a similar result to our in-house-developed SNP-probe 371 and 417 assays. The results of this study indicate that the SNP-probe we designed can be used in the detection of the SARS-CoV-2 Omicron variant.

Comparative Analysis of the Protein Composition of Goat Milk from French Alpine, Nubian, and Creole Breeds and Holstein Friesian Cow Milk: Implications for Early Infant Nutrition.

Simple SummaryGoat’s milk is a food that contains proteins of value for nutrition. The protein profile in the milk of goat breeds is different from that of cow milk, with a lower relative abundance of allergenic proteins. In addition, regardless of the breed, goat milk has beta-casein of type A2 in a more significant proportion than cow milk, which impacts different bioactive peptides hydrolyzed in the milk of the species.Of the diversity of proteins and high digestibility, goat milk will be a food of significant value for infant nutrition. The genetic polymorphisms of milk proteins play an essential role in the different degrees of allergic reactions. This work aimed to identify the proteins and peptides in the composition of goat milk and compare them to those in cow’s milk. The work was performed with goats French Alpine, Nubian, and Creole breeds and Holstein Friesian milking cows at the Universidad Autónoma de Querétaro, Amazcala. We investigated the relative abundance of goat and cow milk protein fractions by SDS-PAGE resolution and the densitometric analysis of gels. The protein alfa-casein was (17.67 ± 0.46) for Creole, (19.18 ± 0.88) French Alpine, (17.35 ± 0.49) Nubian, and (35.92 ± 1.96) Holstein cows. The relative abundance obtained from alfa-casein was statistically different between goats and cows, and this protein was vital because it is a protein related to allergies. On the other hand, the amino acid in position 67 of the beta-casein from three goat breeds is a Proline, so it is assumed that the beta-casein variant of goat milk is an A2-type. The latter has excellent relevance for infant nutrition and differs from cow milk.

Molecular Heterogeneity of Classical and Duarte Galactosemia

For the first time we have conducted molecular-genetic analysis of GALT gene of galactosemia inherited disease in the newborn blood which found at the result of screening conducted maternity houses of Baku city of Azerbaijan Republic. For Azerbaijan people for the first-time identification of substitution of adenine nucleotide with guanine nucleotide in GALT gene position 563 was conducted. It leaded to substitution of glycineamino acid with arginine amino acid in position 188 of mutation protein. Galactosemia is an alteration that alters the way the body processes galactose. It may be due to a deficiency of any of the three enzymes of the galactose catabolic pathway; galactose-1- phosphate urydiltransferase (Gal-1-PUT), galactokinase or urydil diphosphate (UDP)- galactose-4-epimerase. Individuals having galactosemia are intolerant to lactose and galactose in milk. In the lack of proper diagnosis and treatment, this disorder can lead to enlarged liver, cirrhosis, renal failure, cataracts, brain damage and even death. Long term complications can be prevented with early diagnosis following birth and by eliminating lactose and galactose containing nutrients from daily diet.

The Interaction of the AAT Protein with Alpha‐1 Antitrypsin Deficiency and Other Respiratory Disorders

Alpha‐1 antitrypsin (AAT) deficiency is a codominant genetic disease. Ordinarily, the liver makes large amounts of the protein AAT and releases it into the blood in its normal M form. It was hypothesized that if there is a deficiency of the AAT protein in the human body, which is caused by the misfolding of the Z variant of the protein in the liver, then AAT deficiency disorder occurs, which contributes to a multitude of lung and liver diseases. This Z variant results from an encoded misfolded variant where the glutamic acid in position 342 of the protein is replaced by a lysine. Structurally, AAT is a proteinase inhibitor and binds to human leukocyte elastase, which prevents damage to the lower respiratory tract. AAT deficiency is not a disease itself, but is considered a predisposition for the development of a number of diseases, mainly pulmonary emphysema and other chronic respiratory disorders. Through literature review as well as conversing with an expert on AAT protein and deficiency, it was found that the insufficient release of AAT into the blood causes low blood levels and increased susceptibility to respiratory diseases, as the function of the AAT protein is to provide protection to the lungs against such diseases. In its ideal functionality, conditions such as emphysema, cirrhosis, and COVID‐19 are protected against, but with AAT deficiency, the individual is more susceptible to these conditions. Although AAT deficiency is inherited genetically, environmental factors can contribute to the early onset of the disorder. Some examples are the utilization of lung irritants, such as smoking or vaping, as well as irritants in the forms of particles found in coal mining and steel millwork. Avoiding these environmental factors as much as possible can allow for an individual to mitigate symptoms of AAT deficiency for a prolonged period of time. Currently, there is no permanent cure to this disease other than frequent infusions, but there are various treatments under research. Studies have been focused on the geographical reach of AAT, as well as newly discovered and efficient treatment options for AAT deficiency. Researchers have been additionally synthesizing information pertaining to the AAT protein being heavily correlated with COVID‐19, which can aid in the research of similar respiratory diseases.

Knowledge and Awareness of Sickle Cell Anemia: Cross Sectional Study among the General Population in Saudi Arabia

Sickle cell disease (SCD) is a hereditary blood disorder. The red blood cells are sickle in shape by polymerization of the Hemoglobin molecule caused by a β-globin gene defect that leads to Substitution of valine for glutamic acid in position 6 of the β-globin. It's an autosomal recessive inherited hematological disease. In Saudi Arabia, the prevalence of the sickle cell gene in the adult population is (4.2%) for sickle-cell trait and (0.26%) for SCD, with the highest prevalence was found in the eastern region. Consanguineous marriage gave a high incidence of the disease. The aim of the current study was to build interest to Knowledge assessment & awareness of Sickle Cell Anemia (SCA) in different regions in Saudi society and try to spread more information about it. Also, assessing the willingness of society to do premarital testing to prevent genetical diseases.Results: A total number of 400 participants filled in the present study questionnaire. The overall percentage of awareness was 60.16%

The Amino-Proximal Region of the Coat Protein of Cucumber Vein Yellowing Virus (Family Potyviridae) Affects the Infection Process and Whitefly Transmission.

Most plant viruses rely on vector transmission for their spread and specific interactions between vector and virus have evolved to regulate this relationship. The whitefly Bemisia tabaci- transmitted cucumber vein yellowing virus (CVYV; genus Ipomovirus, family Potyviridae) is endemic in the Mediterranean Basin, where it causes significant losses in cucurbit crops. In this study, the role of the coat protein (CP) of CVYV for B. tabaci transmission and plant infection was investigated using a cloned and infectious CVYV cDNA and a collection of point and deletion mutants derived from this clone. Whitefly transmission of CVYV was abolished in a deletion mutant lacking amino acids in position 93–105 of the CP. This deletion mutant caused more severe disease symptoms compared to the cDNA clone representing the wild-type (wt) virus and movement efficiency was likewise affected. Two virus mutants carrying a partially restored CP were transmissible and showed symptoms comparable to the wt virus. Collectively, our data demonstrate that the N-terminus of the CVYV CP is a determinant for transmission by the whitefly vector and is involved in plant infection and symptom expression.

Antiallergic Activity of 6-Deoxy-2-O-methyl-6-(N-hexadecanoyl)amino-l-ascorbic Acid.

Allergy is an excessive immune response to a specific antigen. Type I allergies, such as hay fever and food allergies, have increased significantly in recent years and have become a worldwide problem. We previously reported that an ascorbic acid derivative having palmitoyl and glucosyl groups, 2-O-α-d-glucopyranosyl-6-O-hexadecanoyl-l-ascorbic acid (6-sPalm-AA-2G), showed inhibitory effects on degranulation in vitro and on the passive cutaneous anaphylaxis (PCA) reaction in mice. In this study, several palmitoyl derivatives of ascorbic acid were synthesized and a structure–activity relationship study was performed to discover more potent ascorbic acid derivatives with degranulation inhibitory activity. 6-Deoxy-2-O-methyl-6-(N-hexadecanoyl)amino-l-ascorbic acid (2-Me-6-N-Palm-AA), in which a methyl group was introduced into the hydroxyl group at the C-2 position of ascorbic acid and in which the hydroxyl group at the C-6 position was substituted with an N-palmitoyl group, exhibited much higher inhibitory activity for degranulation in vitro than did 6-sPalm-AA-2G. 2-Me-6-N-Palm-AA strongly inhibit the PCA reaction in mice at lower doses than those of 6-sPalm-AA-2G. These findings suggest that 2-Me-6-N-Palm-AA may be a promising therapeutic candidate for allergic diseases.

A late-onset male Fabry disease patient with somatic mosaicism of a classical GLA mutation: a case report

Fabry disease (FD) is an X-linked lysosomal storage disorder caused by mutations in the α-galactosidase A gene (GLA). Male patients of FD develop early sign and symptoms in childhood or adolescence. However, "de novo somatic mosaicism" is rare and might be developed a relatively mild phenotype despite carrying a classic type. A 34-year-old male patient visited with foamy urine. Renal biopsy findings were consistent with FD. Leukocyte α-galactosidase activity was markedly reduced at 5.3 nmol/hr/mg (normal range, 25-126). Sequence analysis of the patient's GLA gene identified mosaicism for the mutation GLA[NM_000169.2] c.820=/G>C. This mutation results in a substitution of the amino acid in position 274 from glycine to arginine. However, no family members showed FD-related symptoms, and the daughter of the patient was also tested for paternity and was identified as a real biological daughter, but DNA sequence analysis for FD showed no mutations. Based on these results, we diagnosed the patients as de novo mutation with somatic mosaicism. Next generation sequencing turned out that 58% of the readings had the mutated allele in buccal cells, 84% in blood, and 85% in urine, when 100% should be expected in a hemizygous affected male confirming the presence of somatic mosaicisms. The patient has been on treatment for enzyme replacement therapy (agalsidase-β, 1.0 mg/kg biweekly) for past 9 years and has maintained normal renal function (serum creatinine 1.0 mg/dL) with mild albuminuria (123 mg/g Cr). Therefore, this case suggests somatic mosaicism is one of important phenotype modifiers.

An acidic morpholine derivative containing glyceride from thraustochytrid, Aurantiochytrium

A novel acidic morpholine-derivative containing glyceride (M-glyceride) was isolated from the cells of two strains of the thraustochytrid, Aurantiochytrium. The glyceride accounted for approximately 0.1 -0.4% of the lyophilized cells. The glyceride consisted of peaks I (85%) and II (15%). The structures of the intact and acetylated glycerides were elucidated by liquid chromatography-quadrupole time-of-flight chromatograph mass spectrometer (LC–Q/TOF) and NMR spectroscopy. The hydrate type of M-glyceride was detected as a minor component by LC–MS/MS. By 2D-NMR experiments, peaks I of the intact M-glyceride were elucidated as 1,2-didocosapentaenoyl-glyceryl-2′-oxy-3′-oxomorpholino propionic acid, and peak II was estimated 1,2-palmitoyldocosapentaenoyl- and/or 1,2-docosapentaenoylpalmitoyl-glyceryl-2′-oxy-3′-oxomorpholino propionic acid. The double bond position of docosapentaenoic acid was of the ω − 6 type (C22 = 5.ω − 6). The M-glyceride content varied by the cell cycle. The content was 0.4% of lyophilized cells at the mid logarithmic phase, and decreased to 0.1% at the mid stationary phase. When cells were grown in 1.0 µM M-glyceride-containing growth media, cell growth was stimulated to 110% of the control. With 0.1 µM acetyl M-glyceride, stimulation of 113% of the control was observed. Finding morpholine derivatives in biological components is rare, and 2-hydroxy-3-oxomorpholino propionic acid (auranic acid) is a novel morpholine derivative.

A 2D Non-degeneracy Graphical Representation of Protein Sequence and Its Applications

Background: The comparison of the protein sequences is an important research filed in bioinformatics. Many alignment-free methods have been proposed. Objective: In order to mining the more information of the protein sequence, this study focus on a new alignment-free method based on physiochemical properties of amino acids. Methods: Average physiochemical value (Apv) has been defined. For a given protein sequence, a 2D curve was outlined based on Apv and position of the amino acid, and there is not loop and intersection on the curve. According to the curve, the similarity/dissimilarity of the protein sequences can be analyzed. Results and Conclusion: Two groups of protein sequences are taken as examples to illustrate the new methods, the protein sequences can be classified correctly, and the results are highly correlated with that of ClustalW. The new method is simple and effective.

Experimental and density functional theory study of oxygen reduction reaction at liquid-liquid interface by oxidovanadium(IV)-4-methyl salophen complex

The catalytic reduction of oxygen (O2) by using oxidovanadium(IV)-4-methyl salophen (VOL) indicated that this complex can efficiently catalyze this reaction in the presence of ferrocene (Fc) as a weak electron donor at the interface between two immiscible electrolyte solutions (ITIES). The oxidovanadium complex has four positions for transferring proton at interface. Proton affinity (PA), gas phase basicity (GB), and pKa for these positions were calculated by using DFT method. Protonated complex (VOL-OH+) is as an active intermediate specie in this catalytic process. The catalytic oxygen reduction by VOL at liquid-liquid interface was also studied by density functional theory (DFT) computations in order to find the effect of structural parameters on this catalytic reaction. Theoretical studies indicated that the oxygen atom of V=O moiety is the most acidic and the best position for transferring proton and electron to the molecular oxygen. The results showed that the catalytic reaction can proceed by direct interaction between molecular oxygen and most acidic position in the structure of VOL.

Optimized Opioid-Neurotensin Multitarget Peptides: From Design to Structure-Activity Relationship Studies.

Fusion of nonopioid pharmacophores, such as neurotensin, with opioid ligands represents an attractive approach for pain treatment. Herein, the μ-/δ-opioid agonist tetrapeptide H-Dmt-d-Arg-Aba-β-Ala-NH2 (KGOP01) was fused to NT(8-13) analogues. Since the NTS1 receptor has been linked to adverse effects, selective MOR-NTS2 ligands are preferred. Modifications were introduced within the native NT sequence, particularly a β3-homo amino acid in position 8 and Tyr11 substitutions. Combination of β3hArg and Dmt led to peptide 7, a MOR agonist, showing the highest NTS2 affinity described to date (Ki = 3 pM) and good NTS1 affinity (Ki = 4 nM), providing a >1300-fold NTS2 selectivity. The (6-OH)Tic-containing analogue 9 also exhibited high NTS2 affinity (Ki = 1.7 nM), with low NTS1 affinity (Ki = 4.7 μM), resulting in an excellent NTS2 selectivity (>2700). In mice, hybrid 7 produced significant and prolonged antinociception (up to 8 h), as compared to the KGOP01 opioid parent compound.