Artificial Cysteine S‐Glycosylation Induced by Per‐O‐Acetylated Unnatural Monosaccharides during Metabolic Glycan Labeling

Qin, Wei; Qin, Ke; Fan, Xinqi; Peng, Linghang; Hong, Weiyao; Zhu, Yuntao; Lv, Pinou; Du, Yifei; Huang, Rongbing; Han, Meihua; Cheng, Bo; Liu, Yuan; Zhou, Wen; Wang, Chu; Chen, Xing

doi:10.1002/ange.201711710

Cited by 54 publications

(50 citation statements)

References 16 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…While these data are suggestive, formal demonstration of intracellular incorporation requires direct evidence that the probe labels peptidoglycan precursors. More broadly, better characterization of the metabolic fate of probes would increase the precision of conclusions that can be drawn from labeling experiments (26, 27).…”

Section: Introductionmentioning

confidence: 99%

Peptidoglycan precursor synthesis along the sidewall of pole-growing mycobacteria

García‐Heredia¹,

Pohane

Melzer

et al. 2018

Preprint

View full text Add to dashboard Cite

1 2 D-amino acid probes label cell wall peptidoglycan at both the poles and sidewall of pole-growing 3 mycobacteria. Since peptidoglycan assembly along the cell periphery could provide a rapid, 4 growth-independent means by which to edit the cell wall, we sought to clarify the precise 5 metabolic fates of these probes. D-amino acid monopeptides were incorporated into 6 peptidoglycan by L,D-transpeptidase remodeling enzymes to varying extents. Dipeptides were 7 incorporated into cytoplasmic precursors. While dipeptide-marked peptidoglycan synthesis at 8 the poles was associated with cell elongation, synthesis along the periphery was highly 9 responsive to cell wall damage. Our observations suggest a post-expansion role for 1 0 peptidoglycan assembly along the mycobacterial sidewall and provide a conceptual framework 1 1 for understanding cell wall robustness in the face of polar growth.

show abstract

Section: Introductionmentioning

confidence: 99%

Peptidoglycan precursor synthesis along the sidewall of pole-growing mycobacteria

García‐Heredia¹,

Pohane

Melzer

et al. 2018

Preprint

View full text Add to dashboard Cite

show abstract

“…In an Ac4ManNAzand time-dependent manner, we observed biotinylated species in the very high (>10 kilobases) molecular weight (MW) region. It has recently been reported that high doses of azidosugars can produce non-enzymatic protein labeling (14), however, in vitro incubation of total RNA with up to 20 mM Ac4ManNAz did not produce the previously observed biotinylated species on RNAs in the high MW region ( fig. S1A).…”

mentioning

confidence: 79%

Mammalian Y RNAs are modified at discrete guanosine residues with N-glycans

Flynn

Bah

Johnson

et al. 2019

Preprint

View full text Add to dashboard Cite

Glycans modify lipids and proteins to mediate inter-and intramolecular interactions across all domains of life. RNA, another multifaceted biopolymer, is not thought to be a major target of glycosylation. Here, we challenge this view with evidence that mammalian cells use RNA as a third scaffold for glycosylation in the secretory pathway. Using a battery of chemical and biochemical approaches, we find that a select group of small noncoding RNAs including Y RNAs are modified with complex, sialylated N-glycans (glycoRNAs). These glycoRNA are present in multiple cell types and mammalian species, both in cultured cells and in vivo. Finally, we find that RNA glycosylation depends on the canonical N-glycan biosynthetic machinery within the ER/Golgi luminal spaces. Collectively, these findings suggest the existence of a ubiquitous interface of RNA biology and glycobiology suggesting an expanded role for glycosylation beyond canonical lipid and protein scaffolds. MAINGlycans have been shown to regulate a wide array of critical biological processes, ranging from cell-cell contacts to host-pathogen interactions, and even the organization of multicellular organisms(1). In a traditionally adjacent field of study, RNA represents another biopolymer that is central to all known life. While the building blocks of RNA are canonically limited to four bases, post-transcriptional modifications (PTMs) can dramatically elaborate the chemical diversity of RNA, with >100 identified PTMs(2-4). The cellular role for RNA is more complex than that of a simple messenger. For instance, RNAs function as scaffolds, molecular decoys, enzymes, and network regulators across the nucleus and cytosol(5-7). With the exception of a few monosaccharide-based tRNA modifications (8,9), there has been no evidence of a direct interface between these two fields of biology.

show abstract

“…This information may be especially useful in the interpretation of experiments conducted using these important tools, as metabolic engineering with azido sugars has been employed in many studies with various applications, such as the dynamics of O-GlcNAcylation and in vivo imaging of glycosylation events. 3,9 However, despite its straightforward concept, the metabolism of these azido sugars has the potential to pose pitfalls for studies using these tools. Many important questions still need to be answered to better understand and interpret results from MGE, such as whether the non-natural sugars undergo interconversion just like natural monosaccharides, how the change in flux of sugar donors and the different tolerance of glycan-processing enzymes for non-natural sugars will shape the cellular glycome and into which exact positions these non-natural sugars are incorporated.…”

Section: Dynamics Of Non-natural Monosaccharide Incorporation Throughmentioning

confidence: 99%

“…In addition, non-specific background interactions with other reactive species in the proteome often leads to false interpretations. [7][8][9] Several attempts to reduce these nonspecific reactions, which are proposed to be due to free radical mediated bond formation, are reported, but none of the methods were able to eliminate them completely. [10][11][12] Even though recent strategies of using isotopic probes for chemical enrichment and isotopic recoding address the false detection of glycopeptides to a considerable extent, the use of tag-based enrichment can lead to ambiguity due to nonspecific and incomplete reactions.…”

Section: Introductionmentioning

confidence: 99%

Mass spectrometric method for the unambiguous profiling of cellular dynamic glycosylation

Shajahan

Supekar

et al. 2020

Preprint

View full text Add to dashboard Cite

Various biological processes at the cellular level are regulated by glycosylation which is a highly micro-heterogeneous post-translational modification (PTM) on proteins and lipids. The dynamic nature of glycosylation can be studied through bio-orthogonal tagging of metabolically engineered nonnatural sugars into glycan epitopes. However, this approach possesses a significant drawback due to nonspecific background reactions and ambiguity of non-natural sugar metabolism. Here we report a tag-free strategy for their direct detection by glycoproteomics and glycomics using mass spectrometry. The method dramatically simplifies the detection of non-natural functional group bearing monosaccharides installed through promiscuous sialic acid, GalNAc, and GlcNAc biosynthetic pathways. Multistage enrichment of glycoproteins by cellular fractionation, subsequent ZIC-HILIC based glycopeptide enrichment, and a spectral enrichment algorithm for the MS data processing enabled direct detection of nonnatural monosaccharides that are incorporated at low abundance on the N/O-glycopeptides along with their natural counterparts. Our approach allowed the detection of both natural and non-natural sugar bearing glycopeptides, N and O-glycopeptides, differentiation of non-natural monosaccharide types on the glycans and also their incorporation efficiency through quantitation. Through this we could deduce some interconversion of monosaccharides during their processing through glycan salvage pathway and subsequent incorporation into glycan chains. The study of glycosylation dynamics through this method can be conducted in high throughput as few sample processing steps are involved, enabling understanding of glycosylation dynamics under various external stimuli and thereby could bolster the use of metabolic glycan engineering in glycosylation functional studies.

show abstract

Artificial Cysteine S‐Glycosylation Induced by Per‐O‐Acetylated Unnatural Monosaccharides during Metabolic Glycan Labeling

Cited by 54 publications

References 16 publications

Peptidoglycan precursor synthesis along the sidewall of pole-growing mycobacteria

Peptidoglycan precursor synthesis along the sidewall of pole-growing mycobacteria

Mammalian Y RNAs are modified at discrete guanosine residues with N-glycans

Mass spectrometric method for the unambiguous profiling of cellular dynamic glycosylation

Contact Info

Product

Resources

About