Analysis of Synthetic Monodisperse Polysaccharides by Wide Mass Range Ultrahigh-Resolution MALDI Mass Spectrometry

Nicolardi, Simone; Joseph, A. Abragam; Zhu, Qian; Shen, Zhengnan; Pardo-Vargas, Alonso; Chiodo, Fabrizio; Molinaro, Antonio; Silipo, Alba; Burgt, Yuri E. M. van der; Yu, Biao; Seeberger, Peter H.; Wuhrer, Manfred

doi:10.1021/acs.analchem.1c00239

Cited by 22 publications

(18 citation statements)

References 76 publications

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“…The purification and characterization of the final glycans might be the major bottlenecks in the automated synthesis of glycans, in which practical purification techniques and advanced characterization tools are highly desirable to overcome the solubility issues and characterization problems of highly polar and ultra‐long polysaccharides. [ 81‐83 ] With the rapid progress of automated glycan synthesis, the structure‐ property relationships of a large variety of glycans will be revealed to promote the applications of glycans as diagnostic tools, vaccine candidates, therapeutic agents, and carbohydrate materials.…”

Section: Discussionmentioning

confidence: 99%

Automated Chemical Solid‐Phase Synthesis of Glycans

Yang

2022

Chin. J. Chem.

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Automated chemical solid-phase synthesis is an automation platform for rapid and reliable synthesis of glycans. Since the seminal work of Automated Glycan Assembly (AGA) disclosed by Seeberger in 2001, AGA has evolved from a proof-of-concept to a robust and reliable technology for streamlined production of various types of glycans. Through more than 20 years of unceasing efforts, the major breakthroughs in AGA including linkers, approved building blocks, and synthesizers have been acquired, and numerous influential achievements have been made in complex glycan synthesis. In addition, the HPLC-assisted automated synthesis emerges as a promising automation platform to access glycans. In this review, we highlight the key advances in the field of automated chemical solid-phase synthesis, especially in AGA. The synthesis of representative glycans based on AGA is also described.

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Section: Discussionmentioning

confidence: 99%

Automated Chemical Solid‐Phase Synthesis of Glycans

Yang

2022

Chin. J. Chem.

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“…The purification and characterization of long polysaccharides poses an additional challenge, especially for ionic polysaccharides [ 398 , 441 ]. As the size of synthetic polysaccharides approaches the size of macromolecules, the implementation of novel analytical and purifications techniques becomes necessary [ 283 ].…”

Section: Discussionmentioning

confidence: 99%

“…As synthetic polymannosides have now reached the size of macromolecules, new challenges are arising in terms of purification and characterization. Gel permeation chromatography (GPC) and advanced high-resolution MS analysis are being developed to address the need of these new available compounds [ 278 , 283 ].…”

Section: Reviewmentioning

confidence: 99%

Progress and challenges in the synthesis of sequence controlled polysaccharides

Fittolani¹,

Tyrikos‐Ergas²,

Vargová³

et al. 2021

Beilstein J. Org. Chem.

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The sequence, length and substitution of a polysaccharide influence its physical and biological properties. Thus, sequence controlled polysaccharides are important targets to establish structure–properties correlations. Polymerization techniques and enzymatic methods have been optimized to obtain samples with well-defined substitution patterns and narrow molecular weight distribution. Chemical synthesis has granted access to polysaccharides with full control over the length. Here, we review the progress towards the synthesis of well-defined polysaccharides. For each class of polysaccharides, we discuss the available synthetic approaches and their current limitations.

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“… 38 − 40 The combination of MALDI-ISD with ultrahigh-resolution FT-ICR MS enables the detailed analysis of both protein and carbohydrate fragment ions over an extended m / z range. 39 , 41 , 42 Interestingly, MALDI-ISD of intact glycoproteins has only provided protein sequence information through the analysis of protein backbone fragment ions. To date, glycan information has only been obtained through the detection of glycosylated protein fragments (or their MS/MS analysis) and not from the direct detection of glycan fragments that were generated from in-source fragmentation of the glycan moieties from the proteins.…”

Section: Introductionmentioning

confidence: 99%

“…MALDI-in-source decay (ISD) is a fragmentation technique that can be applied to both intact proteins and polysaccharides. − The combination of MALDI-ISD with ultrahigh-resolution FT-ICR MS enables the detailed analysis of both protein and carbohydrate fragment ions over an extended m / z range. ,, Interestingly, MALDI-ISD of intact glycoproteins has only provided protein sequence information through the analysis of protein backbone fragment ions. To date, glycan information has only been obtained through the detection of glycosylated protein fragments (or their MS/MS analysis) and not from the direct detection of glycan fragments that were generated from in-source fragmentation of the glycan moieties from the proteins. , MALDI analysis is fast and robust and does not require upfront separation techniques as often needed for ESI MS. MALDI is particularly suitable for high-throughput analyses and features short measurement times.…”

Section: Introductionmentioning

confidence: 99%

Glycan and Protein Analysis of Glycoengineered Bacterial E. coli Vaccines by MALDI-in-Source Decay FT-ICR Mass Spectrometry

et al. 2022

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Bacterial glycoconjugate vaccines have a major role in preventing microbial infections. Immunogenic bacterial glycans, such as O-antigen polysaccharides, can be recombinantly expressed and combined with specific carrier proteins to produce effective vaccines. O-Antigen polysaccharides are typically polydisperse, and carrier proteins can have multiple glycosylation sites. Consequently, recombinant glycoconjugate vaccines have a high structural heterogeneity, making their characterization challenging. Since development and quality control processes rely on such characterization, novel strategies are needed for faster and informative analysis. Here, we present a novel approach employing minimal sample preparation and ultrahigh-resolution mass spectrometry analysis for protein terminal sequencing and characterization of the oligosaccharide repeat units of bacterial glycoconjugate vaccines. Three glycoconjugate vaccine candidates, obtained from the bioconjugation of the O-antigen polysaccharides from E. coli serotypes O2, O6A, and O25B with the genetically detoxified exotoxin A from Pseudomonas aeruginosa , were analyzed by MALDI-in-source decay (ISD) FT-ICR MS. Protein and glycan ISD fragment ions were selectively detected using 1,5-diaminonaphtalene and a 2,5-dihydroxybenzoic acid/2-hydroxy-5-methoxybenzoic acid mixture (super-DHB) as a MALDI matrix, respectively. The analysis of protein fragments required the absence of salts in the samples, while the presence of salt was key for the detection of sodiated glycan fragments. MS/MS analysis of O-antigen ISD fragments allowed for the detection of specific repeat unit signatures. The developed strategy requires minute sample amounts, avoids the use of chemical derivatizations, and comes with minimal hands-on time allowing for fast corroboration of key structural features of bacterial glycoconjugate vaccines during early- and late-stage development.

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Analysis of Synthetic Monodisperse Polysaccharides by Wide Mass Range Ultrahigh-Resolution MALDI Mass Spectrometry

Cited by 22 publications

References 76 publications

Automated Chemical Solid‐Phase Synthesis of Glycans

Automated Chemical Solid‐Phase Synthesis of Glycans

Progress and challenges in the synthesis of sequence controlled polysaccharides

Glycan and Protein Analysis of Glycoengineered Bacterial E. coli Vaccines by MALDI-in-Source Decay FT-ICR Mass Spectrometry

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