Synthesis of small protein domains by automated flow chemistry

Ferentzi, Kristóf; Nagy-Fazekas, Dóra; Farkas, Viktor; Perczel, András

doi:10.1039/d3re00324h

Cited by 5 publications

(3 citation statements)

References 53 publications

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“…a All amino acids were coupled using our in-house-developed SPF method and apparatus. However, the last N-terminal amino acids were varied [22][23][24]. b Conditions used after coupling the Fmoc-Thr(ΨPro)-OH (#1-#5) or Fmoc-Thr( t Bu)-OH (#6) residue to see the effect of temperature after coupling.…”

Section: The 5mer* Problemmentioning

confidence: 99%

“…The use of Xxx(ΨPro) as monomeric building blocks has been the subject of only a few attempts [19][20][21][22]. In light of these considerations, the choice of Xxx(ΨPro) over the expensive Yyy-Xxx(ΨPro) dipeptides would be a desirable approach, using our in-house-developed smart peptide chemistry in flow method or SPF for short [22][23][24] (Figure 2). SPF is not just a device within HPLC modules or just a synthetic protocol; it represents a complex, optimized approach that takes into account synthetic time, reagent quantities, and environmental impact.…”

Section: Introductionmentioning

confidence: 99%

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Unveiling the Oxazolidine Character of Pseudoproline Derivatives by Automated Flow Peptide Chemistry

Szaniszló,

Csámpai,

Horváth

et al. 2024

IJMS

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Pseudoproline derivatives such as Thr(ΨPro)-OH are commonly used in peptide synthesis to reduce the likelihood of peptide aggregation and to prevent aspartimide (Asi) formation during the synthesis process. In this study, we investigate notable by-products such as aspartimide formation and an imine derivative of the Thr(ΨPro) moiety observed in flow peptide chemistry synthesis. To gain insight into the formation of these unexpected by-products, we design a series of experiments. Furthermore, we demonstrate the oxazolidine character of the pseudoproline moiety and provide plausible mechanisms for the two-way ring opening of oxazolidine leading to these by-products. In addition, we present evidence that Asi formation appears to be catalyzed by the presence of the pseudoproline moiety. These observed side reactions are attributed to elevated temperature and pressure; therefore, caution is advised when using ΨPro derivatives under such harsh conditions. In addition, we propose a solution whereby thermodynamically controlled Asi formation can be kinetically prevented.

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Section: The 5mer* Problemmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Unveiling the Oxazolidine Character of Pseudoproline Derivatives by Automated Flow Peptide Chemistry

Szaniszló,

Csámpai,

Horváth

et al. 2024

IJMS

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“…Hardware development would also be crucial for efficient experimentation in chemical protein synthesis. Currently, there have been advancements in microware-assisted peptide synthesis − and flow chemistry-based peptide synthesis. − It is anticipated that future developments will lead to the creation of robots capable of automating peptide synthesis, peptide ligation, and analyzing reactions using techniques like high-performance liquid chromatography (HPLC) and mass spectrometry (MS). These robots would also be able to automatically conduct product purification and protein folding.…”

Section: Outlook: Opportunity and Challengesmentioning

confidence: 99%

Chemical Synthesis of Human Proteoforms and Application in Biomedicine

Ai,

Pan,

Liu

2024

ACS Cent. Sci.

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Limited understanding of human proteoforms with complex posttranslational modifications and the underlying mechanisms poses a major obstacle to research on human health and disease. This Outlook discusses opportunities and challenges of de novo chemical protein synthesis in human proteoform studies. Our analysis suggests that to develop a comprehensive, robust, and cost-effective methodology for chemical synthesis of various human proteoforms, new chemistries of the following types need to be developed: (1) easy-to-use peptide ligation chemistries allowing more efficient de novo synthesis of protein structural domains, (2) robust temporary structural support strategies for ligation and folding of challenging targets, and (3) efficient transpeptidative protein domain−domain ligation methods for multidomain proteins. Our analysis also indicates that accurate chemical synthesis of human proteoforms can be applied to the following aspects of biomedical research: (1) dissection and reconstitution of the proteoform interaction networks, (2) structural mechanism elucidation and functional analysis of human proteoform complexes, and (3) development and evaluation of drugs targeting human proteoforms. Overall, we suggest that through integrating chemical protein synthesis with in vivo functional analysis, mechanistic biochemistry, and drug development, synthetic chemistry would play a pivotal role in human proteoform research and facilitate the development of precision diagnostics and therapeutics.

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