NMR Spectroscopic Characterization of the C‐Mannose Conformation in a Thrombospondin Repeat Using a Selective Labeling Approach

Jonker, Hendrik R. A.; Saxena, Krishna; Shcherbakova, Aleksandra; Tiemann, Birgit; Bakker, Hans; Schwalbe, Harald

doi:10.1002/anie.202009489

Cited by 12 publications

(12 citation statements)

References 44 publications

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“…The C-mannosides 12a−f all adopted the typical 4 C 1 conformation, while the pyranose of the glycopeptides 12g−i all preferred the 1 C 4 conformation. This provides further evidence that tryptophan plays a defining role in driving a shift in the pyranose's preferred conformation from 4 C 1 to 1 C 4 , though the mechanism underlying this change is not immediately obvious.…”

Section: ■ Results and Discussionmentioning

confidence: 99%

“…Tryptophan C-mannosylation is the only known form of protein C-glycosylation. It involves the formation of a C–C bond between C-2 of L -tryptophan and the anomeric position of d -mannopyranose to give 2-(α- d -mannopyranosyl)- L -tryptophan or Trp(Man) 1 . , This C-mannoside adopts the unusual 1 C 4 conformation rather than the 4 C 1 conformer preferred by most other mannosides. − The modification is found on a diverse array of proteins, including those with a thrombospondin repeat (TSR) domain, type-I cytokine receptors, RNases, , lipoprotein lyase, glycoside hydrolases, glycosyltransferases, siglecs, and viral glycoproteins, yet its biological functions remain poorly characterized.…”

Section: Introductionmentioning

confidence: 99%

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Synthesis of C-Mannosylated Glycopeptides Enabled by Ni-Catalyzed Photoreductive Cross-Coupling Reactions

Mao

Shah

et al. 2021

J. Am. Chem. Soc.

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The biological functions of tryptophan C-mannosylation are poorly understood, in part, due to a dearth of methods for preparing pure glycopeptides and glycoproteins with this modification. To address this issue, efficient and scalable methods are required for installing this protein modification. Here, we describe unique Ni-catalyzed cross-coupling conditions that utilize photocatalysis or a Hantzsch ester photoreductant to couple glycosyl halides with (hetero)aryl bromides, thereby enabling the α-C-mannosylation of 2-bromo-tryptophan, peptides thereof, and (hetero)aryl bromides more generally. We also report that 2-(α-Dmannopyranosyl)-L-tryptophan undergoes facile anomerization in the presence of acid: something that must be considered when preparing and handling peptides with this modification. These developments enabled the first automated solid-phase peptide syntheses of C-mannosylated glycopeptides, which we used to map the epitope of an antibody, as well as providing the first verified synthesis of Carmo-HrTH-I, a C-mannosylated insect hormone. To complement this approach, we also performed late-stage tryptophan C-mannosylation on a diverse array of peptides, demonstrating the broad scope and utility of this methodology for preparing glycopeptides.

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Section: ■ Results and Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Synthesis of C-Mannosylated Glycopeptides Enabled by Ni-Catalyzed Photoreductive Cross-Coupling Reactions

Mao

Shah

et al. 2021

J. Am. Chem. Soc.

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“…In X-ray structures, C-mannose residues have been observed in both 4 C 1 and 1 C 4 conformations. , However, NMR studies show a preference for the unusual 1 C 4 ring shape on RNase 2 and within a TSR. , The binding of mannose to BC2L-A is possible only in a 4 C 1 conformation via the two calcium ions. To be able to bind to BC2L-A, the C-mannose thus needs to adopt a 4 C 1 conformation.…”

Section: Discussionmentioning

confidence: 99%

A Bacterial Mannose Binding Lectin as a Tool for the Enrichment of C- and O-Mannosylated Peptides

et al. 2022

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Mass spectrometry (MS) easily detects C-mannosylated peptides from purified proteins but not from complex biological samples. Enrichment of specific glycopeptides by lectin affinity prior to MS analysis has been widely applied to support glycopeptide identification but was until now not available for C-mannosylated peptides. Here, we used the α-mannose-specific Burkholderia cenocepacia lectin A (BC2L-A) and show that, in addition to its previously demonstrated high-mannose N-glycan binding capability, this lectin is able to retain C- and O-mannosylated peptides. Besides testing binding abilities to standard peptides, we applied BC2L-A affinity to enrich C-mannosylated peptides from complex samples of tryptic digests of HEK293 and MCF10A whole cell extracts, which led to the identification of novel C-mannosylation sites. In conclusion, BC2L-A enabled specific enrichment of C- and O-mannosylated peptides and might have superior properties over other mannose binding lectins for this purpose.

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“…Besides chemical synthesis, mannosylated proteins were reportedly expressed in Drosophila Schneider 2 (S2) cells [ 44 ]. Mutation of mannose phosphate isomerase (MPI) by CRISPR-CAS can abolish mannose generation activity from glucose ( Figure 4 ).…”

Section: Structural and Synthetic Biology Of Protein C -Mannosylation And C -Man-trpmentioning

confidence: 99%

Protein C-Mannosylation and C-Mannosyl Tryptophan in Chemical Biology and Medicine

et al. 2021

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C-Mannosylation is a post-translational modification of proteins in the endoplasmic reticulum. Monomeric α-mannose is attached to specific Trp residues at the first Trp in the Trp-x-x-Trp/Cys (W-x-x-W/C) motif of substrate proteins, by the action of C-mannosyltransferases, DPY19-related gene products. The acceptor substrate proteins are included in the thrombospondin type I repeat (TSR) superfamily, cytokine receptor type I family, and others. Previous studies demonstrated that C-mannosylation plays critical roles in the folding, sorting, and/or secretion of substrate proteins. A C-mannosylation-defective gene mutation was identified in humans as the disease-associated variant affecting a C-mannosylation motif of W-x-x-W of ADAMTSL1, which suggests the involvement of defects in protein C-mannosylation in human diseases such as developmental glaucoma, myopia, and/or retinal defects. On the other hand, monomeric C-mannosyl Trp (C-Man-Trp), a deduced degradation product of C-mannosylated proteins, occurs in cells and extracellular fluids. Several studies showed that the level of C-Man-Trp is upregulated in blood of patients with renal dysfunction, suggesting that the metabolism of C-Man-Trp may be involved in human kidney diseases. Together, protein C-mannosylation is considered to play important roles in the biosynthesis and functions of substrate proteins, and the altered regulation of protein C-manosylation may be involved in the pathophysiology of human diseases. In this review, we consider the biochemical and biomedical knowledge of protein C-mannosylation and C-Man-Trp, and introduce recent studies concerning their significance in biology and medicine.

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NMR Spectroscopic Characterization of the C‐Mannose Conformation in a Thrombospondin Repeat Using a Selective Labeling Approach

Cited by 12 publications

References 44 publications

Synthesis of C-Mannosylated Glycopeptides Enabled by Ni-Catalyzed Photoreductive Cross-Coupling Reactions

Synthesis of C-Mannosylated Glycopeptides Enabled by Ni-Catalyzed Photoreductive Cross-Coupling Reactions

A Bacterial Mannose Binding Lectin as a Tool for the Enrichment of C- and O-Mannosylated Peptides

Protein C-Mannosylation and C-Mannosyl Tryptophan in Chemical Biology and Medicine

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