scholarly journals Oxyamine Linkers for the Protecting Group Free Synthesis of Glycoconjugates

2021 ◽  
Author(s):  
◽  
Jaimé Caitlin Hill

<p>Glycoconjugates, such as glycolipids and glycoproteins, are involved in a variety of cellular functions including cell-to-cell signalling and carbohydrate-protein recognition. Accordingly, glycoconjugates play important roles in health and disease and are promising new leads as carbohydrate-based therapeutics. However, for the development of glycoconjugates to study biological processes, or for the use of these adducts as therapeutics, the glycan needs to be conjugated to the carrier molecule or scaffold of choice. Many procedures for the conjugation of glycans involve lengthy protecting group strategies that install the aglycone at the start of glycan total synthesis and are therefore unsuitable for naturally derived sugars. Other glycan conjugation strategies can affect the integrity of the reducing end sugar or lead to adducts where the reducing end sugar adopts the ring-opened rather than the ring-closed form. N,O-Dialkyl oxyamine linkers, however, can be attached to the free reducing end of sugars in a single step without the need for protecting groups. This thesis therefore explores the synthesis and application of oxyamine linkers for the synthesis of glycoconjugates.  First, the synthesis of an O-alkyl-N-methyl oxyamine linker (“Type A”) containing an amine at its terminus was improved by reducing the number of synthetic steps from six to four and by increasing the overall yield from 8% to 38%. This oxyamine linker was then conjugated to GlcNAc in 83% yield. The hydrolytic stability of this glycosyloxyamine was then compared to that of the analogous N-alkyl-O-methyl glycosyloxyamine (“Type B”). The stability of the two types of glycosyloxyamines has never been directly compared. Accordingly, it was not known whether the difference in substitution pattern between the two linkers affects their hydrolytic stability. To this end, the hydrolysis rates of the GlcNAc conjugated linkers were assessed at various pH values, glycoconjugate concentrations and buffer concentrations. In all instances, the “Type B” glycoside was found to have marginally better kinetic stability, while the “Type A” glycoside had marginally better thermodynamic stability, but overall, these differences were negligible. The pKa of the conjugate acid of these glycosyloxyamines was also determined to provide insight into the mechanism of hydrolysis. By considering this data, along with the observation that the rate of hydrolysis of these glycoconjugates increases with increasing buffer concentration, it was proposed that the hydrolysis of the oxyamines occurs via general acid catalysis at pH 4-6.  A novel dithiol functionalised oxyamine linker was also designed and synthesised for the multivalent display of glycans on gold nanoparticles. With the successful attachment of this thiol linker to GlcNAc, the monomer unit of chitin, this work has paved the way for the future syntheses of chitin-functionalised gold nanoparticles. Such chitinfunctionalised AuNPs can be used to assess chitin’s ability to invoke the asthma allergic immune response, thereby bringing the possibility of an anti-asthma vaccine a step closer to fruition.</p>

2021 ◽  
Author(s):  
◽  
Jaimé Caitlin Hill

<p>Glycoconjugates, such as glycolipids and glycoproteins, are involved in a variety of cellular functions including cell-to-cell signalling and carbohydrate-protein recognition. Accordingly, glycoconjugates play important roles in health and disease and are promising new leads as carbohydrate-based therapeutics. However, for the development of glycoconjugates to study biological processes, or for the use of these adducts as therapeutics, the glycan needs to be conjugated to the carrier molecule or scaffold of choice. Many procedures for the conjugation of glycans involve lengthy protecting group strategies that install the aglycone at the start of glycan total synthesis and are therefore unsuitable for naturally derived sugars. Other glycan conjugation strategies can affect the integrity of the reducing end sugar or lead to adducts where the reducing end sugar adopts the ring-opened rather than the ring-closed form. N,O-Dialkyl oxyamine linkers, however, can be attached to the free reducing end of sugars in a single step without the need for protecting groups. This thesis therefore explores the synthesis and application of oxyamine linkers for the synthesis of glycoconjugates.  First, the synthesis of an O-alkyl-N-methyl oxyamine linker (“Type A”) containing an amine at its terminus was improved by reducing the number of synthetic steps from six to four and by increasing the overall yield from 8% to 38%. This oxyamine linker was then conjugated to GlcNAc in 83% yield. The hydrolytic stability of this glycosyloxyamine was then compared to that of the analogous N-alkyl-O-methyl glycosyloxyamine (“Type B”). The stability of the two types of glycosyloxyamines has never been directly compared. Accordingly, it was not known whether the difference in substitution pattern between the two linkers affects their hydrolytic stability. To this end, the hydrolysis rates of the GlcNAc conjugated linkers were assessed at various pH values, glycoconjugate concentrations and buffer concentrations. In all instances, the “Type B” glycoside was found to have marginally better kinetic stability, while the “Type A” glycoside had marginally better thermodynamic stability, but overall, these differences were negligible. The pKa of the conjugate acid of these glycosyloxyamines was also determined to provide insight into the mechanism of hydrolysis. By considering this data, along with the observation that the rate of hydrolysis of these glycoconjugates increases with increasing buffer concentration, it was proposed that the hydrolysis of the oxyamines occurs via general acid catalysis at pH 4-6.  A novel dithiol functionalised oxyamine linker was also designed and synthesised for the multivalent display of glycans on gold nanoparticles. With the successful attachment of this thiol linker to GlcNAc, the monomer unit of chitin, this work has paved the way for the future syntheses of chitin-functionalised gold nanoparticles. Such chitinfunctionalised AuNPs can be used to assess chitin’s ability to invoke the asthma allergic immune response, thereby bringing the possibility of an anti-asthma vaccine a step closer to fruition.</p>


2021 ◽  
Vol 16 (1) ◽  
pp. 13-20
Author(s):  
Trilianty Lestarisa ◽  
Heny Arwati ◽  
Yoes Prijatna Dachlan ◽  
Soedjajadi Keman ◽  
Din Syafruddin

Background: Plasmodium vivax is transmitted most across the country of Indonesia. The country has set out a malaria elimination program by 2030. The information on genetic diversity of malarial parasites relates to malaria transmission in an endemic area may provide the information that can help the malaria control program to achieve the target. Therefore, the purpose of this study was to determine the genetic diversity of the Pvmsp-1 gene in Central Kalimantan Province. Materials and Methods: Samples were 140 of archived Giemsa-stained blood smear and rapid detection test. Samples were divided into the indigenous and migrant populations. After confirmation by single-step PCR, only P. vivax and mixed infection samples were amplified to nested PCR for genotyping of Pvmsp-1 allelic variation in segments F1, F2, and F3. Results: Genotyping of 23 PCR positive samples resulted in 13 genotypes. In segment F1, three allelic variants type A containing subtype A1 (1,050 bp), A2 (350 bp), A3 (150 bp), and type B (100 bp). In segment F2, mono genotypes were detected as variant type A (1,050 bp) and type B3 (150 bp), multiple genotypes were detected as type B containing subtype B1 (250 bp), B2 (200 bp), and B3 (150bp). In segment F3, three allelic variants generated from four mono genotypes were type A (350 bp), type B (300 bp), and two type C (250 bp). Conclusion: The low allelic variation of Pvmsp-1 gene may reflect the actual situation of the low malaria endemic status of the study sites.


Author(s):  
S. Fujinaga ◽  
K. Maruyama ◽  
C.W. Williams ◽  
K. Sekhri ◽  
L. Dmochowski

Yumoto and Dmochowski (Cancer Res.27, 2098 (1967)) reported the presence of mature and immature type C leukemia virus particles in leukemic organs and tissues such as lymph nodes, spleen, thymus, liver, and kidneys of SJL/J strain mice with Hodgki's-like disease or reticulum cell neoplasm (type B). In an attempt to ascertain the possibility that this neoplasia may be of viral origin, experiments with induction and transmission of this neoplasm were carried out using cell-free extracts of leukemic organs from an SJL/J strain mouse with spontaneous disease.It has been possible to induce the disease in low-leukemia BALB/c and C3HZB strain mice and serially transfer the neoplasia by cell-free extracts of leukemic organs of these mice. Histological examination revealed the neoplasia to be of either reticulum cell-type A or type B. Serial transfer is now in its fifth passage. In addition leukemic spleen from another SJL/J strain mouse with spontaneous reticulum cell neoplasm (type A) was set up in tissue culture and is now in its 141st serial passage in vitro. Preliminary results indicate that cell-free material of 39th tissue culture passage can reproduce neoplasia in BALB/c mice.


1992 ◽  
Vol 68 (03) ◽  
pp. 297-300 ◽  
Author(s):  
Monica Galli ◽  
Paul Comfurius ◽  
Tiziano Barbui ◽  
Robert F A Zwaal ◽  
Edouard M Bevers

SummaryPlasmas of 16 patients positive for both IgG anticardiolipin (aCL) antibodies and lupus anticoagulant (LA) antibodies were subjected to adsorption with liposomes containing cardiolipin. In 5 of these plasmas both the anticardiolipin and the anticoagulant activities were co-sedimented with the liposomes in a dose-dependent manner, whereas in the remaining cases only the anticardiolipin activity could be removed by the liposomes, leaving the anticoagulant activity (LA) in the supernatant plasma. aCL antibodies purified from the first 5 plasmas were defined as aCL-type A, while the term aCL-type B was used for antibodies in the other 11 plasmas, from which 2 were selected for this study.Prolongation of the dRVVT was produced by affinity-purified aCL-type A antibodies in plasma of human as well as animal (bovine, rat and goat) origin. aCL-type B antibodies were found to be devoid of anticoagulant activity, while the corresponding supernatants containing LA IgG produced prolongation of the dRVVT only in human plasma.These anticoagulant activities of aCL-type A and of LA IgG's were subsequently evaluated in human plasma depleted of β2-glycoprotein I (β2-GPI), a protein which was previously shown to be essential in the binding of aCL antibodies to anionic phospholipids. Prolongation of the dRVVT by aCL-type A antibodies was abolished using β2-GPI deficient plasma, but could be restored upon addition of β2-GPI. In contrast, LA IgG caused prolongation of the dRVVT irrespective of the presence or absence of β2-GPI.Since β2-GPI binds to negatively-charged phospholipids and impedes the conversion of prothrombin by the factor Xa/Va enzyme complex (Nimpf et al., Biochim Biophys Acta 1986; 884: 142–9), comparison was made of the effect of aCL-type A and aCL-type B antibodies on the rate of thrombin formation in the presence and absence of β2-GPI. This was measured in a system containing highly purified coagulation factors Xa, Va and prothrombin and lipid vesicles composed of 20 mole% phosphatidylserine and 80 mole% phosphatidylcholine. No inhibition on the rate of thrombin formation was observed with both types of aCL antibodies when either β2-GPI or the lipid vesicles were omitted. Addition of β2-GPI to the prothrombinase assay in the presence of lipid vesicles causes a time-dependent inhibition which was not affected by the presence of aCL-type B or non-specific IgG. In contrast, the presence of aCL-type A antibodies dramatically increased the anticoagulant effect of β2-GPI. These data indicate that the anticoagulant activity of aCL-type A antibodies in plasma is mediated by β2-GPI.


2013 ◽  
Vol 1 (1) ◽  
pp. 10
Author(s):  
Noar Muda Satyawan ◽  
Shelly Tutupoho ◽  
Yusli Wardiatno ◽  
Makoto Tsuchiya

Erosion rate on corals due to activities of other biota is called bioerosion. The rock-boring urchin, Echinometra mathaei, when it is abundant, plays a significant role in benthic ecosystems, including biological processes like coral erosion. During feeding, E. mathaei erodes calcium carbonate besides grazing on algae living on coral, so it plays an important role in both organic and inorganic carbons in coral reefs. The urchin E. mathaei actively feeds during the night time (nocturnal grazer). Although in Okinawa four types (A-D) of the urchin exist, the research only focused on the types A and B. Type A of E. mathaei produced 0.44951 g feces per day on average while type B produced 0.38030 g feces per day. CaCO3 analysis in feces and gut contents showed bioerosion rate of E. mathaei type A was 0.64492 g/individu/day, and 0.54436 g/individu/day in type B. There were no significant differences in bioerosion impact of E. mathaei type A and B© Laju erosi pada karang yang disebabkan oleh biota, dikenal dengan bioerosi. Bulu babi jenis Echinometra mathaei, ketika melimpah, menjadi sangat berpengaruh terhadap ekosistem bentik termasuk proses biologi seperti erosi karang. Selama aktivitas makan, E. mathaei menggerus kalsium karbonat dalam proporsi yang besar di samping alga yang tumbuh menempel pada karang sehingga memiliki peran penting dalam siklus karbon organik dan anorganik di ekosistem terumbu karang. Bulu babi E. mathaei aktif mencari makan pada malam hari (nocturnal grazer). Meskipun di Okinanawa ada 4 tipe (A-D), pada eksperimen kali ini memfokuskan pada tipe A dan B saja. Tipe A E. mathaei rata-rata memproduksi 0,44951 g feses/hari dan tipe B memproduksi 0,38030 g feses/hari. Berdasarkan analisis CaCO3 yang dilakukan pada feses dan isi lambung, laju bioerosi yang disebabkan oleh E. mathaei tipe A sebesar 0,64492 g/individu/hari sedangkan tipe B sebesar 0,54436 g/individu/hari. Tidak terdapat perbedaan dampak bioerosi yang signifikan antara E. mathaei tipe A dan B©


2020 ◽  
Vol 22 (Supplement_2) ◽  
pp. ii41-ii41
Author(s):  
Junjie Zhen ◽  
Lei Wen ◽  
Shaoqun Li ◽  
Mingyao Lai ◽  
Changguo Shan ◽  
...  

Abstract BACKGROUND According to EANO-ESMO clinical practice guidelines, the MRI findings of LM are divided into 4 types, namely linear enhancement (type A), nodular enhancement (type B), linear combined with nodular enhancement (type C), and sign of hydrocephalus (type D). METHODS The MRI features of brain and spinal cord in patients diagnosed with NSCLC-LM in Guangdong Sanjiu Brain Hospital from 2010 until 2019 were investigated, and then were classified into 4 types. The imaging features were analyzed. RESULTS A total of 80 patients were enrolled in the study. The median age of the patients was 53.5 years old, and the median time from the initial diagnosis to the confirmed diagnosis of LM was 11.6 months. The results of enhanced MRI examination of the brain in 79 cases showed that the number of cases with enhancements of type A, B, C and D were 50 (63.3%), 0, 26 (32.9%) and 3 (3.8%), respectively, and that LM with metastases to the brain parenchyma was found in 42 cases (53.2%). The results of enhanced MRI examination of spinal cord in 59 cases showed that there were only enhancements of type A and C in 40 cases (67.8%) and 3 cases (5.0%), and no enhancement sign in the other 16 cases (27.2%). CONCLUSION MRI examination of brain and spinal cord will improve the detection rate of LM. The MRI features of NSCLC-LM in real world are mainly characterized by the linear enhancements of brain and spinal cord, followed by linear combined with nodular enhancement. The enhancements of type B and type D are rare in clinic. Almost half of the patients have LM and metastases to the brain parenchyma. Therefore, the differentiation of tumor metastases is needed to be paid attention to for the early diagnosis and the formulation of reasonable treatment plans.


BMC Genomics ◽  
2021 ◽  
Vol 22 (1) ◽  
Author(s):  
Hüseyin Can ◽  
Sedef Erkunt Alak ◽  
Ahmet Efe Köseoğlu ◽  
Umut Şahar ◽  
Berna Bostanbaş ◽  
...  

Abstract Background Cytidine monophospho-n-acetylneuraminic acid hydroxylase (CMAH) gene associated with blood groups in cats encodes CMAH enzyme that converts Neu5Ac to Neu5Gc. Although variations in CMAH gene of pedigree cats have been revealed, the presence/lack of them in non-pedigree stray cats is unknown. Therefore, the present study aimed to investigate the variations in CMAH gene and the quantity of Neu5Ac and Neu5Gc on erythrocytes of non-pedigree stray cats (n:12) living in İzmir, Turkey. Also, the frequency of blood types was determined in 76 stray cats including 12 cats that were used for CMAH and Neu5A/Neu5Gc analysis. Results In total, 14 SNPs were detected in 5’UTR as well as in exon 2, 4, 9, 10, 11 and 12 of CMAH gene. Among these SNPs, -495 C > T in 5’UTR was detected for the first time as heterozygous in type A and AB cats, and homozygous and heterozygous in type B cats. The remaining 13 that have been detected in previous studies were also found as homozygous or heterozygous. Both Neu5Gc and Neu5Ac were detected in type A and AB cats. In type B cats, only Neu5Ac was detected. Among two type AB cats, the level of Neu5Ac was found higher in cat carrying heterozygous form (T/C) of 1392T > C. The prevalence of type B cats (67.1 %) was higher than others. Conclusions The presence of a new SNP as well as previous SNPs indicates that more variations can be found in stray cats with a more comprehensive study in the future. Also, the high prevalence of type B cats demonstrates the possible risk of neonatal isoerythrolysis among stray cats living in İzmir, Turkey.


2019 ◽  
Vol 40 (Supplement_1) ◽  
Author(s):  
T Yamaguchi ◽  
M Nakai ◽  
Y Sumita ◽  
Y Miyamoto ◽  
H Matsuda ◽  
...  

Abstract Background Despite recent advances in diagnosis and management, the mortality of acute aortic dissection (AAD) remains high. Purpose This study aims to develop quality indicators (QIs) for the management of AAD, and to evaluate the associations between QIs and outcomes of AAD in a Japanese nationwide administrative database. Methods A total of 18,348 patients suffered from AAD (Type A: 10,131, Type B: 8,217) in the Japanese Registry of All Cardiac and Vascular Diseases database between 2012 and 2015 were studied. A systematic review was performed to establish initial index items for QIs. Evaluation was performed through the expert consensus meeting using a Delphi method. Associations between developed QIs and the mortality were determined by multivariate mixed logistic regression analyses. Results A total of nine QIs (five structural and four processatic) were developed. Achievements of developed QIs (High: 7–9, Middle: 4–6, Low: 0–3) were significantly associated with lower in-hospital mortality even after adjustment for covariates in both type A (Middle: odds ratio [OR], 0.257; 95% confidence interval [CI], 0.211–0.312; P<0.001; High: OR, 0.064; 95% CI, 0.047–0.086; P<0.001 vs. Low) and type B (Middle: OR, 0.447; 95% CI, 0.338–0.590; P<0.001; High: OR, 0.128; 95% CI, 0.077–0.215; P<0.001 vs. Low). Additionally, achievements of structural and processatic QIs were consistently associated with reduced in-hospital mortality. QIs and in-hospital mortality Conclusions Developed QIs for AAD management were significantly associated with lower in-hospital mortality. Evaluation of each hospital's management with QIs could be helpful to equalize quality of treatment and to fill the evidence-to-practice gaps in the real-world treatment.


2019 ◽  
Vol 201 (19) ◽  
Author(s):  
Surashree S. Kulkarni ◽  
Joseph J. Johnston ◽  
Yongtao Zhu ◽  
Zachary T. Hying ◽  
Mark J. McBride

ABSTRACTFlavobacterium johnsoniaeSprB moves rapidly along the cell surface, resulting in gliding motility. SprB secretion requires the type IX secretion system (T9SS). Proteins secreted by the T9SS typically have conserved C-terminal domains (CTDs) belonging to the type A CTD or type B CTD family. Attachment of 70- to 100-amino-acid type A CTDs to a foreign protein allows its secretion. Type B CTDs are common but have received little attention. Secretion of the foreign protein superfolder green fluorescent protein (sfGFP) fused to regions spanning the SprB type B CTD (sfGFP-CTDSprB) was analyzed. CTDs of 218 amino acids or longer resulted in secretion of sfGFP, whereas a 149-amino-acid region did not. Some sfGFP was secreted in soluble form, whereas the rest was attached on the cell surface. Surface-attached sfGFP was rapidly propelled along the cell, suggesting productive interaction with the motility machinery. This did not result in rapid cell movement, which apparently requires additional regions of SprB. Secretion of sfGFP-CTDSprBrequired coexpression withsprF, which lies downstream ofsprB. SprF is similar in sequence toPorphyromonas gingivalisPorP. MostF. johnsoniaegenes encoding proteins with type B CTDs lie immediately upstream ofporP/sprF-like genes. sfGFP was fused to the type B CTD from one such protein (Fjoh_3952). This resulted in secretion of sfGFP only when it was coexpressed with its cognate PorP/SprF-like protein. These results highlight the need for extended regions of type B CTDs and for coexpression with the appropriate PorP/SprF-like protein for efficient secretion and cell surface localization of cargo proteins.IMPORTANCETheF. johnsoniaegliding motility adhesin SprB is delivered to the cell surface by the type IX secretion system (T9SS) and is rapidly propelled along the cell by the motility machinery. How this 6,497-amino-acid protein interacts with the secretion and motility machines is not known. Fusion of the C-terminal 218 amino acids of SprB to a foreign cargo protein resulted in its secretion, attachment to the cell surface, and rapid movement by the motility machinery. Efficient secretion of SprB required coexpression with the outer membrane protein SprF. Secreted proteins that have sequence similarity to SprB in their C-terminal regions are common in the phylumBacteroidetesand may have roles in adhesion, motility, and virulence.


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