RESUMO
A primary pathology of Alzheimer's disease (AD) is amyloid ß (Aß) deposition in brain parenchyma and blood vessels, the latter being called cerebral amyloid angiopathy (CAA). Parenchymal amyloid plaques presumably originate from neuronal Aß precursor protein (APP). Although vascular amyloid deposits' origins remain unclear, endothelial APP expression in APP knock-in mice was recently shown to expand CAA pathology, highlighting endothelial APP's importance. Furthermore, two types of endothelial APP-highly O-glycosylated APP and hypo-O-glycosylated APP-have been biochemically identified, but only the former is cleaved for Aß production, indicating the critical relationship between APP O-glycosylation and processing. Here, we analyzed APP glycosylation and its intracellular trafficking in neurons and endothelial cells. Although protein glycosylation is generally believed to precede cell surface trafficking, which was true for neuronal APP, we unexpectedly observed that hypo-O-glycosylated APP is externalized to the endothelial cell surface and transported back to the Golgi apparatus, where it then acquires additional O-glycans. Knockdown of genes encoding enzymes initiating APP O-glycosylation significantly reduced Aß production, suggesting this non-classical glycosylation pathway contributes to CAA pathology and is a novel therapeutic target.
Assuntos
Acetilgalactosamina , Doença de Alzheimer , Peptídeos beta-Amiloides , Precursor de Proteína beta-Amiloide , Angiopatia Amiloide Cerebral , Glicosilação , Animais , Camundongos , Doença de Alzheimer/complicações , Doença de Alzheimer/metabolismo , Doença de Alzheimer/patologia , Peptídeos beta-Amiloides/biossíntese , Peptídeos beta-Amiloides/química , Peptídeos beta-Amiloides/metabolismo , Precursor de Proteína beta-Amiloide/química , Precursor de Proteína beta-Amiloide/metabolismo , Angiopatia Amiloide Cerebral/complicações , Angiopatia Amiloide Cerebral/metabolismo , Angiopatia Amiloide Cerebral/patologia , Células Endoteliais/metabolismo , Transporte Proteico , Neurônios/metabolismo , Complexo de Golgi/metabolismo , Acetilgalactosamina/metabolismoRESUMO
KEY MESSAGE: Genotyping-by-sequencing of 723 worldwide cucumber genetic resources revealed that cucumbers were dispersed eastward via at least three distinct routes, one to Southeast Asia and two from different directions to East Asia. The cucumber (Cucumis sativus) is an economically important vegetable crop cultivated and consumed worldwide. Despite its popularity, the manner in which cucumbers were dispersed from their origin in South Asia to the rest of the world, particularly to the east, remains a mystery due to the lack of written records. In this study, we performed genotyping-by-sequencing (GBS) on 723 worldwide cucumber accessions, mainly deposited in the Japanese National Agriculture and Food Research Organization (NARO) Genebank, to characterize their genetic diversity, relationships, and population structure. Analyses based on over 60,000 genome-wide single-nucleotide polymorphisms identified by GBS revealed clear genetic differentiation between Southeast and East Asian populations, suggesting that they reached their respective region independently, not progressively. A deeper investigation of the East Asian population identified two subpopulations with different fruit characteristics, supporting the traditional classification of East Asian cucumbers into two types thought to have been introduced by independent routes. Finally, we developed a core collection of 100 accessions representing at least 93.2% of the genetic diversity present in the entire collection. The genetic relationships and population structure, their associations with geographic distribution and phenotypic traits, and the core collection presented in this study are valuable resources for elucidating the dispersal history and promoting the efficient use and management of genetic resources for research and breeding in cucumber.
Assuntos
Cucumis sativus , Polimorfismo de Nucleotídeo Único , Cucumis sativus/genética , Genética Populacional , Genótipo , Variação Genética , Ásia OrientalRESUMO
L-Arabinofuranosides with ß-linkages are present in several plant molecules, such as arabinogalactan proteins (AGPs), extensin, arabinan, and rhamnogalacturonan-II. We previously characterized a ß-L-arabinofuranosidase from Bifidobacterium longum subsp. longum JCM 1217, Bll1HypBA1, which was found to belong to the glycoside hydrolase (GH) family 127. This strain encodes two GH127 genes and two GH146 genes. In the present study, we characterized a GH146 ß-L-arabinofuranosidase, Bll3HypBA1 (BLLJ_1848), which was found to constitute a gene cluster with AGP-degrading enzymes. This recombinant enzyme degraded AGPs and arabinan, which contain Araf-ß1,3-Araf structures. In addition, the recombinant enzyme hydrolyzed oligosaccharides containing Araf-ß1,3-Araf structures but not those containing Araf-ß1,2-Araf and Araf-ß1,5-Araf structures. The crystal structures of Bll3HypBA1 were determined at resolutions up to 1.7 Å. The monomeric structure of Bll3HypBA1 comprised a catalytic (α/α)6 barrel and two ß-sandwich domains. A hairpin structure with two ß-strands was observed in Bll3HypBA1, to extend from a ß-sandwich domain and partially cover the active site. The active site contains a Zn2+ ion coordinated by Cys3-Glu and exhibits structural conservation of the GH127 cysteine glycosidase Bll1HypBA1. This is the first study to report on a ß1,3-specific ß-L-arabinofuranosidase. KEY POINTS: ⢠ß1,3-l-Arabinofuranose residues are present in arabinogalactan proteins and arabinans as a terminal sugar. ⢠ß-l-Arabinofuranosidases are widely present in intestinal bacteria. ⢠Bll3HypBA1 is the first enzyme characterized as a ß1,3-linkage-specific ß-l-arabinofuranosidase.
Assuntos
Bifidobacterium , Glicosídeo Hidrolases , Catálise , CisteínaRESUMO
Rare sugars are known for their ability to suppress postprandial blood glucose levels. Therefore, oligosaccharides and disaccharides derived from rare sugars could potentially serve as functional sweeteners. A disaccharide [α-d-allopyranosyl-(1â2)-ß-d-psicofuranoside] mimicking sucrose was synthesized from rare monosaccharides D-allose and D-psicose. Glycosylation using the intermolecular aglycon delivery (IAD) method was employed to selectively form 1,2-cis α-glycosidic linkages of the allopyranose residues. Moreover, ß-selective psicofuranosylation was performed using a psicofuranosyl acceptor with 1,3,4,6-tetra-O-benzoyl groups. This is the first report on the synthesis of non-reducing disaccharides comprising only rare d-sugars by IAD using protected ketose as a unique acceptor; additionally, this approach is expected to be applicable to the synthesis of functional sweeteners.
Assuntos
Dissacarídeos , Frutose , Glucose , Sacarose , Dissacarídeos/química , Dissacarídeos/síntese química , Sacarose/química , Glicosilação , Edulcorantes/químicaRESUMO
Methods for producing drugs directly at the cancer site, particularly using bioorthogonal metal catalysts, are being explored to mitigate the side effects of therapy. Albumin-based artificial metalloenzymes (ArMs) catalyze reactions in living mice while protecting the catalyst in the hydrophobic pocket. Here, we describe the in situ preparation and application of biocompatible tumor-targeting ArMs using circulating albumin, which is abundant in the bloodstream. The ArM was formed using blood albumin through the intravenous injection of ruthenium conjugated with an albumin-binding ligand; the tumor-targeting unit was conjugated to the ArM using its catalytic activity, and the ArM was transported to the cancer site. The delivered ArM catalyzed a second tagging reaction of the proapoptotic peptide on the cancer surface, successfully suppressing cancer proliferation. This approach, which efficiently leveraged the persisting reactivity twice in vivo, holds promise for future in vivo metal-catalyzed drug synthesis utilizing endogenous albumin.
RESUMO
In plant cell walls, the hydroxyproline-rich glycoproteins (HRGPs) such as extensin contain oligoarabinofuranoside linked to a hydroxyproline (Hyp) residue. The mature arabinooligosaccharide was revealed to be a tetrasaccharide (α-l-Araf-(1â3)-ß-l-Araf-(1â2)-ß-l-Araf-(1â2)-ß-l-Araf, l-Araf4 ), whose linkages are targets of the bifidobacterial and Xanthomonas arabinooligosaccharide-degrading enzymes. The l-Araf4 motif was cleaved by GH43 α-l-arabinofuranosidase (Arafase) and converted to an l-Araf3 -linked structure. The latter is then cleaved by GH121 ß-l-arabinobiosidase (HypBA2), producing ß-l-Araf-(1â2)-l-Ara (ß-l-arabinobiose) and mono-ß-l-Araf linked to the HRGP backbone. In bifidobacteria, the ß-l-arabinobiose is then hydrolyzed by GH127 ß-l-Arafase (Bll1HypBA1), a mechanistically unique cysteine glycosidase. We recently identified the distantly related homologue from Xanthomonas euvesicatoria as GH146 ß-l-Arafase along with paralogues from Bifidobacterium longum, one of which, Bll4HypBA1 (BLLJ_0089), can degrade l-Araf1 -Hyp in a similar way to that of GH146. As the chemical synthesis of the extensin hydrophilic motif 1 a, which possesses three distinct linkages that connect four oligoAraf residues [Hyp(l-Arafn ) (n=4, 3, 1)], was achieved previously, we precisely monitored the step-wise enzymatic cleavage of 1 a in addition to that of potato lectin. The results unequivocally revealed that this enzyme specifically degrades the Hyp(l-Araf1 ) motif.
Assuntos
Bifidobacterium , Glicosídeo Hidrolases , Bifidobacterium/metabolismo , Hidroxiprolina , Glicosídeo Hidrolases/metabolismo , GlicoproteínasRESUMO
To uncover population structure, phylogenetic relationship, and diversity in melons along the famous Silk Road, a seed size measurement and a phylogenetic analysis using five chloroplast genome markers, 17 RAPD markers and 11 SSR markers were conducted for 87 Kazakh melon accessions with reference accessions. Kazakh melon accessions had large seed with exception of two accessions of weedy melon, Group Agrestis, and consisted of three cytoplasm types, of which Ib-1/-2 and Ib-3 were dominant in Kazakhstan and nearby areas such as northwestern China, Central Asia and Russia. Molecular phylogeny showed that two unique genetic groups, STIa-2 with Ib-1/-2 cytoplasm and STIa-1 with Ib-3 cytoplasm, and one admixed group, STIAD combined with STIa and STIb, were prevalent across all Kazakh melon groups. STIAD melons that phylogenetically overlapped with STIa-1 and STIa-2 melons were frequent in the eastern Silk Road region, including Kazakhstan. Evidently, a small population contributed to melon development and variation in the eastern Silk Road. Conscious preservation of fruit traits specific to Kazakh melon groups is thought to play a role in the conservation of Kazakh melon genetic variation during melon production, where hybrid progenies were generated through open pollination.
RESUMO
Numerous genetic resources of major crops have been introduced from around the world and deposited in Japanese National Agriculture and Food Research Organization (NARO) Genebank. Understanding their genetic variation and selecting a representative subset ("core collection") are essential for optimal management and efficient use of genetic resources. In this study, we conducted genotyping-by-sequencing (GBS) to characterize the genetic relationships and population structure in 755 accessions of melon genetic resources. The GBS identified 39,324 single-nucleotide polymorphisms (SNPs) that are distributed throughout the melon genome with high density (one SNP/10.6 kb). The phylogenetic relationships and population structure inferred using this SNP dataset are highly associated with the cytoplasm type and geographical origin. Our results strongly support the recent hypothesis that cultivated melon was established in Africa and India through multiple independent domestication events. Finally, we constructed a World Melon Core Collection that covers at least 82% of the genetic diversity and has a wide range of geographical origins and fruit morphology. The genome-wide SNP dataset, phylogenetic relationships, population structure, and the core collection provided in this study should largely contribute to genetic research, breeding, and genetic resource preservation in melon.
RESUMO
Controlling the stereoselectivity of 1,2-cis glycosylation is one of the most challenging tasks in the chemical synthesis of glycans. There are various 1,2-cis glycosides in nature, such as α-glucoside and ß-mannoside in glycoproteins, glycolipids, proteoglycans, microbial polysaccharides, and bioactive natural products. In the structure of polysaccharides such as α-glucan, 1,2-cis α-glucosides were found to be the major linkage between the glucopyranosides. Various regioisomeric linkages, 1â3, 1â4, and 1â6 for the backbone structure, and 1â2/3/4/6 for branching in the polysaccharide as well as in the oligosaccharides were identified. To achieve highly stereoselective 1,2-cis glycosylation, including α-glucosylation, a number of strategies using inter- and intra-molecular methodologies have been explored. Recently, Zn salt-mediated cis glycosylation has been developed and applied to the synthesis of various 1,2-cis linkages, such as α-glucoside and ß-mannoside, via the 1,2-cis glycosylation pathway and ß-galactoside 1,4/6-cis induction. Furthermore, the synthesis of various structures of α-glucans has been achieved using the recent progressive stereoselective 1,2-cis glycosylation reactions. In this review, recent advances in stereoselective 1,2-cis glycosylation, particularly focused on α-glucosylation, and their applications in the construction of linear and branched α-glucans are summarized.
Assuntos
Glucanos , Polissacarídeos , Glicosilação , Glucanos/química , Glucosídeos , Manosídeos , EstereoisomerismoRESUMO
Over the past decades, 2(5H)-furanone derivatives have been extensively studied because of their promising ability to prevent the biofilm formation by various pathogenic bacteria. Here, we report the synthesis of a series of optically active sulfur-containing 2(5H)-furanone derivatives and characterize their biological activity. Novel thioethers were obtained by an interaction of stereochemically pure 5-(l)-menthyloxy- or 5-(l)-bornyloxy-2(5H)-furanones with aromatic thiols under basic conditions. Subsequent thioethers oxidation by an excess of hydrogen peroxide in acetic acid resulted in the formation of the corresponding chiral 2(5H)-furanone sulfones. The structure of synthesized compounds was confirmed by IR and NMR spectroscopy, HRMS, and single crystal X-ray diffraction. The leading compound, 26, possessing the sulfonyl group and l-borneol moiety, exhibited the prominent activity against Staphylococcus aureus and Bacillus subtilis with MICs of 8 µg/mL. Furthermore, at concentrations of 0.4-0.5 µg/mL, the sulfone 26 increased two-fold the efficacy of aminoglycosides gentamicin and amikacin against S. aureus. The treatment of the model-infected skin wound in the rat with a combination of gentamicin and sulfone 26 speeded up the bacterial decontamination and improved the healing of the wound. The presented results provide valuable new insights into the chemistry of 2(5H)-furanone derivatives and associated biological activities.
Assuntos
Bactérias , Staphylococcus aureus , Ratos , Animais , Antibacterianos/farmacologia , Antibacterianos/química , Amicacina , Gentamicinas , Furanos/químicaRESUMO
Fructooligosaccharides and their anhydrides are widely used as health-promoting foods and prebiotics. Various enzymes acting on ß-D-fructofuranosyl linkages of natural fructan polymers have been used to produce functional compounds. However, enzymes that hydrolyze and form α-D-fructofuranosyl linkages have been less studied. Here, we identified the BBDE_2040 gene product from Bifidobacterium dentium (α-D-fructofuranosidase and difructose dianhydride I synthase/hydrolase from Bifidobacterium dentium [αFFase1]) as an enzyme with α-D-fructofuranosidase and α-D-arabinofuranosidase activities and an anomer-retaining manner. αFFase1 is not homologous with any known enzymes, suggesting that it is a member of a novel glycoside hydrolase family. When caramelized fructose sugar was incubated with αFFase1, conversions of ß-D-Frup-(2â1)-α-D-Fruf to α-D-Fruf-1,2':2,1'-ß-D-Frup (diheterolevulosan II) and ß-D-Fruf-(2â1)-α-D-Fruf (inulobiose) to α-D-Fruf-1,2':2,1'-ß-D-Fruf (difructose dianhydride I [DFA I]) were observed. The reaction equilibrium between inulobiose and DFA I was biased toward the latter (1:9) to promote the intramolecular dehydrating condensation reaction. Thus, we named this enzyme DFA I synthase/hydrolase. The crystal structures of αFFase1 in complex with ß-D-Fruf and ß-D-Araf were determined at the resolutions of up to 1.76 Å. Modeling of a DFA I molecule in the active site and mutational analysis also identified critical residues for catalysis and substrate binding. The hexameric structure of αFFase1 revealed the connection of the catalytic pocket to a large internal cavity via a channel. Molecular dynamics analysis implied stable binding of DFA I and inulobiose to the active site with surrounding water molecules. Taken together, these results establish DFA I synthase/hydrolase as a member of a new glycoside hydrolase family (GH172).
Assuntos
Proteínas de Bactérias/química , Bifidobacterium/enzimologia , Glicosídeo Hidrolases/química , Modelos Moleculares , Oligossacarídeos/química , Cristalografia por Raios X , Glicosídeo Hidrolases/classificaçãoRESUMO
Gum arabic is an arabinogalactan protein (AGP) that is effective as a prebiotic for the growth of bifidobacteria in the human intestine. We recently identified a key enzyme in the glycoside hydrolase (GH) family 39, 3-O-α-d-galactosyl-α-l-arabinofuranosidase (GAfase), for the assimilation of gum arabic AGP in Bifidobacterium longum subsp. longum. The enzyme released α-d-Galp-(1â3)-l-Ara and ß-l-Arap-(1â3)-l-Ara from gum arabic AGP and facilitated the action of other enzymes for degrading the AGP backbone and modified sugar. In this study, we identified an α-l-arabinofuranosidase (BlArafE; encoded by BLLJ_1850), a multidomain enzyme with both GH43_22 and GH43_34 catalytic domains, as a critical enzyme for the degradation of modified α-l-arabinofuranosides in gum arabic AGP. Site-directed mutagenesis approaches revealed that the α1,3/α1,4-Araf double-substituted gum arabic AGP side chain was initially degraded by the GH43_22 domain and subsequently cleaved by the GH43_34 domain to release α1,3-Araf and α1,4-Araf residues, respectively. Furthermore, we revealed that a tetrasaccharide, α-l-Rhap-(1â4)-ß-d-GlcpA-(1â6)-ß-d-Galp-(1â6)-d-Gal, was a limited degradative oligosaccharide in the gum arabic AGP fermentation of B. longum subsp. longum JCM7052. The oligosaccharide was produced from gum arabic AGP by the cooperative action of the three cell surface-anchoring enzymes, GAfase, exo-ß1,3-galactanase (Bl1,3Gal), and BlArafE, on B. longum subsp. longum JCM7052. Furthermore, the tetrasaccharide was utilized by the commensal bacteria. IMPORTANCE Terminal galactose residues of the side chain of gum arabic arabinogalactan protein (AGP) are mainly substituted by α1,3/α1,4-linked Araf and ß1,6-linked α-l-Rhap-(1â4)-ß-d-GlcpA residues. This study found a multidomain BlArafE with GH43_22 and GH43_34 catalytic domains showing cooperative action for degrading α1,3/α1,4-linked Araf of the side chain of gum arabic AGP. In particular, the GH43_34 domain of BlArafE was a novel α-l-arabinofuranosidase for cleaving the α1,4-Araf linkage of terminal galactose. α-l-Rhap-(1â4)-ß-d-GlcpA-(1â6)-ß-d-Galp-(1â6)-d-Gal tetrasaccharide was released from gum arabic AGP by the cooperative action of GAfase, GH43_24 exo-ß-1,3-galactanase (Bl1,3Gal), and BlArafE and remained after B. longum subsp. longum JCM7052 culture. Furthermore, in vitro assimilation test of the remaining oligosaccharide using Bacteroides species revealed that cross-feeding may occur from bifidobacteria to other taxonomic groups in the gut.
Assuntos
Bifidobacterium longum , Bifidobacterium longum/metabolismo , Galactanos/metabolismo , Glicosídeo Hidrolases/metabolismo , Goma Arábica , Humanos , Oligossacarídeos/químicaRESUMO
Recently, the development of abiotic metal-mediated drug delivery has been significant growth in the fields of anticancer approach and biomedical application. However, the intrinsic toxicity of abiotic metal catalysts makes in vivo use difficult. Our group developed a system of cancer-targeting albumin-based artificial metalloenyzmes (ArMs) capable of performing localized drug synthesis and selective tagging therapy in vivo for cancer therapy. The toxicity of the system at higher concentrations was investigated in vitro and in vivo in the study to demonstrate its safety for potential application in clinical trials. In cell-based experiments, the study revealed that the cytotoxicity of metal catalysts anchored within the binding cavity of the cancer-targeting ArMs could be significantly reduced compared to free-in-solution metal catalysts. Moreover, the in vivo data demonstrated that the cancer-targeting ArMs did not cause considerable damage in organs or change in the hematological parameters in a single-dose (160 mg/Kg) toxicity study in rats. Therefore, the system is safe, highlighting that it could be used in clinical trials for cancer treatment.
Assuntos
Metaloproteínas , Neoplasias , Albuminas , Animais , Catálise , Metaloproteínas/metabolismo , Neoplasias/tratamento farmacológico , RatosRESUMO
Methyl ß-l-arabinofuranosyl-(1 â 2)-, -(1 â 3)-, and -(1 â 5)-α-l-arabinofuranosides have been stereoselectively synthesized through 2-naphthylmethyl ether-mediated intramolecular aglycon delivery (NAP-IAD), whose ß-linkages were confirmed by NMR analysis on the 3JH1-H2 coupling constant and 13C chemical shift of C1. The NAP-IAD approach was simply extended for the synthesis of trisaccharide motifs possessing ß-l-arabinofuranosyl-(1 â 5)-l-arabinofuranosyl non-reducing terminal structure with the branched ß-l-arabinofuranosyl-(1 â 5)-[α-l-arabinofuranosyl-(1 â 3)]-α-l-arabinofuranosyl and the liner ß-l-arabinofuranosyl-(1 â 5)-ß-l-arabinofuranosyl-(1 â 5)-ß-l-arabinofuranosyl structures in olive arabinan and dinoflagellate polyethers, respectively. The results on the substrate specificity of a bifidobacterial ß-l-arabinofuranosidase HypBA1 using the regioisomers indicated that HypBA1 could hydrolyze all three linkages however behaved clearly less active to ß-(1 â 5)-linked disaccharide than other two regioisomers including the proposed natural degradation product, ß-(1 â 2)-linked one from plant extracellular matrix such as extensin. On the other hand, Xanthomonas XeHypBA1 was found to hydrolyze all three disaccharides as the substrate with higher specificity to ß-(1 â 2)-linkage than bifidobacterial HypBA1.
Assuntos
Dissacarídeos , Glicosídeo Hidrolases , Glicosídeo Hidrolases/metabolismo , Especificidade por SubstratoRESUMO
To understand the precise mechanism of the glycoside hydrolase (GH) family 127, a cysteine ß-l-arabinofuranosidase (Arafase) - HypBA1 - has been isolated from Bifidobacterium longum in the human Gut microbiota, and the design and synthesis of the mechanism-based inhibitors such as l-Araf-haloacetamides have been carried out. The α-l-Araf-azide derivative was used as the monoglycosylamine equivalent to afford the l-Araf-chloroacetamides (α/ß-1-Cl) as well as bromoacetamides (α/ß-1-Br) in highly stereoselective manner through Staudinger reaction followed by amide formation with/without anomerization. Against HypBA1, the probes 1, especially in the case of α/ß-1-Br inhibited the hydrolysis. Conformational implications of these observations are discussed in this manuscript. Additional examinations using l-Araf-azides (α/ß-5) resulted in further mechanistic observations of the GH127/146 cysteine glycosidases, including the hydrolysis of ß-5 as the substrate and oxidative inhibition by α-5 using the GH127 homologue.
RESUMO
Both pyruvylation and sialylation onto the terminus of oligosaccharides of N-glycoproteins seem to be structurally and functionally similar with a property of conferring negative charge. However, detailed molecular characteristics of pyruvylation and sialylation in vivo were elusive. Here, to investigate an effect of terminal pyruvylation to N-glycan on in vivo biodistribution and kinetics, we prepared human serum albumin (HSA) modified with pyruvylated N-glycan (PvG), conjugated with HiLyte Fluor 750 (FL750-PvGHSA). In vivo imaging by using FL750-PvGHSA revealed that terminally pyruvylated N-glycoalbumin was excreted like sialylated N-glycoalbumin, suggesting that pyruvylation mimics sialylation in in vivo biodistribution and kinetics of N-glycoproteins. Terminal pyruvylation onto N-glycans can be a potential tool for a novel glycoengineering strategy.
Assuntos
Oligossacarídeos , Polissacarídeos , Albuminas , Glicoproteínas/metabolismo , Humanos , Cinética , Distribuição TecidualRESUMO
Neoglycoconjugates mimicking natural compounds and possessing a variety of biological functions are very successful tools for researchers to understand the general mechanisms of many biological processes in living organisms. These substances are characterized by high biotolerance and specificity, with low toxicity. Due to the difficult isolation of individual glycoclusters from biological objects, special interest has been directed toward synthetic analogs. This review is mainly focused on the one-pot, double-click methodology (containing alkyne-azide click cycloaddition with the following 6π-azaelectrocyclization reactions) used in the synthesis of N-glycoconjugates. Homogeneous (including one type of biantennary N-glycan fragments) and heterogeneous (containing two to four types of biantennary N-glycan fragments) glycoclusters on albumin were synthesized via this strategy. A series of cell-, tissue- and animal-based experiments proved glycoclusters to be a very promising class of targeted delivery systems. Depending on the oligosaccharide units combined in the cluster, their amount, and arrangement relative to one another, conjugates can recognize various cells, including cancer cells, with high selectivity. These results open new perspectives for affected tissue visualization and treatment.
Assuntos
Glicoconjugados/síntese química , Polissacarídeos/química , Albumina Sérica Humana/química , Animais , Catálise , Técnicas de Química Sintética , Química Click , Glicoconjugados/química , Humanos , Lectinas/química , Camundongos , Modelos Moleculares , Conformação Molecular , Estrutura Molecular , Relação Estrutura-AtividadeRESUMO
Metabolic oxidation of pyrrolizidine alkaloids (PAs) from herbal and dietary supplements by cytochrome P450 produces dehydro-PAs (DHPs), which leads to toxicities. A highly reactive cation species generated from the active pyrrole ring of DHPs readily reacts with various cellular components, causing hepatotoxicity and cytotoxicity. Inspired by PA-induced hepatic damage, we developed a therapeutic approach based on a cyclization precursor that can be transformed into a synthetic DHP under physiological conditions through gold-catalyzed 5-endo-dig cyclization using a gold-based artificial metalloenzyme (ArM) instead of through metabolic oxidation by cytochrome P450. In cell-based assays, the synthesis of the DHP by a cancer-targeting glycosylated gold-based ArM substantially suppressed cell growth of the targeted cancer cells without causing cytotoxicity to untargeted cells, highlighting the potential of the strategy to be used therapeutically in vivo.
Assuntos
Metaloproteínas , Alcaloides de Pirrolizidina , Alcaloides de Pirrolizidina/toxicidade , Sistema Enzimático do Citocromo P-450 , Pirróis/metabolismo , OuroRESUMO
BACKGROUND: Despite reports of a link between human papillomavirus (HPV) infection and mechanistic target of rapamycin (mTOR) signaling activation, the role of the mTOR pathway, especially raptor and rictor, in HPV-related head and neck cancer is still unclear. The aim of the present study was to elucidate the role of the mTOR pathway in HPV-related oropharyngeal squamous cell carcinoma (OPSCC). METHODS: The present study involved two strategies. The first was to investigate the activity of mTOR and mTOR-related complexes in high-risk HPV-positive (UM-SCC47 and CaSki) and HPV-negative (SCC-4 and SAS) cancer cell lines. The second was to elucidate mTOR complex expression in 80 oropharyngeal cancer tissues and to examine the relationship between mTOR complex expression and survival in patients with OPSCC. RESULTS: The UM-SCC47 and CaSki cell lines showed high gene and protein expression of raptor. They also exhibited G1/S and G2/M phase cell cycle arrest following 24 h incubation with 6 µM temsirolimus, a rapamycin analog, and temsirolimus administration inhibited their growth. HPV-related OPSCC samples showed high gene and protein expression of raptor and rictor compared with HPV-unrelated OPSCC. In addition, HPV-related OPSCC patients with high raptor and rictor expression tended to have a worse prognosis than those with low or medium expression. CONCLUSIONS: These results suggest that raptor and rictor have important roles in HPV-related OPSCC and that temsirolimus is a potential therapeutic agent for patients with HPV-related OPSCC. This is the first report to reveal the overexpression of raptor and rictor in HPV-related OPSCC.
Assuntos
Biomarcadores Tumorais/metabolismo , Carcinoma de Células Escamosas/patologia , Neoplasias Orofaríngeas/patologia , Papillomaviridae/isolamento & purificação , Infecções por Papillomavirus/complicações , Proteína Companheira de mTOR Insensível à Rapamicina/metabolismo , Proteína Regulatória Associada a mTOR/metabolismo , Apoptose , Biomarcadores Tumorais/genética , Carcinoma de Células Escamosas/epidemiologia , Carcinoma de Células Escamosas/metabolismo , Carcinoma de Células Escamosas/virologia , Ciclo Celular , Movimento Celular , Proliferação de Células , Feminino , Regulação Neoplásica da Expressão Gênica , Humanos , Japão/epidemiologia , Masculino , Pessoa de Meia-Idade , Neoplasias Orofaríngeas/epidemiologia , Neoplasias Orofaríngeas/metabolismo , Neoplasias Orofaríngeas/virologia , Infecções por Papillomavirus/virologia , Prognóstico , Taxa de Sobrevida , Células Tumorais CultivadasRESUMO
N-alkyl unsaturated imines derived from acrolein, a toxin produced during oxidative stress, and biogenic alkyl amines occur naturally and are considered biologically relevant compounds. However, despite the recent conceptual and technological advances in organic synthesis, research on the new reactivity of these compounds is lacking. This personal account discusses research on the reactivity that has been overlooked in acrolein imines, including the discovery of new methods to synthesize biologically active compounds, the determination of new functions of relevant imines and their precursors, i. e., aldehydes and amines, and the application of these methods for clinical diagnosis.