Biology subjects

Zong, G.

Publications and source records attributed to Zong, G..

3 recordsLinked to original sources

bioRxiv2023

Ghost fragment ions in structure and site-specific glycoproteomics analysis

Mass spectrometry (MS) can unlock crucial insights into the intricate world of glycosylation analysis. Despite its immense potential, the qualitative and quantitative analysis of isobaric glycopeptide structures remains one of the most daunting hurdles in the field of glycoproteomics. The ability to distinguish between these complex glycan structures poses a significant challenge, hindering our ability to accurately measure and understand the role of glycoproteins in biological systems. A few recent publications described the use of collision energy (CE) modulation to improve structural elucidation, especially for qualitative purposes. Different linkages of glycan units usually demonstrate different stabilities under CID/HCD fragmentation conditions. Fragmentation of the glycan moiety produces low molecular weight ions (oxonium ions) that can serve as a structure-specific signature for specific glycan moieties, however, specificity of these fragments has never been examined closely. Here, we investigated fragmentation specificity using synthetic stable isotope-labelled glycopeptide standards. These standards were isotopically labelled at the reducing terminal GlcNAc, which allowed us to resolve fragments produced by oligomannose core moiety and fragments generated from outer antennary structures. Our research identified the potential for false positive structure assignments due to the occurrence of "Ghost" fragments resulting from single glyco unit rearrangement or mannose core fragmentation within the collision cell. To mitigate this issue, we have established a minimal intensity threshold for these fragments to prevent the misidentification of structure-specific fragments in glycoproteomics analysis. Our findings provide a crucial step forward in the quest for more accurate and reliable glycoproteomics measurements. Graphical abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=142 SRC="FIGDIR/small/541150v1_ufig1.gif" ALT="Figure 1"> View larger version (22K): org.highwire.dtl.DTLVardef@1a5bb21org.highwire.dtl.DTLVardef@1a09b5aorg.highwire.dtl.DTLVardef@137504borg.highwire.dtl.DTLVardef@78831e_HPS_FORMAT_FIGEXP M_FIG C_FIG

biochemistry↗

bioRxiv2022

LC-MS/MS-PRM Quantification of IgG glycoforms using stable isotope labeled IgG1 Fc glycopeptide standard

Targeted quantification of proteins is a standard methodology with broad utility, but targeted quantification of glycoproteins has not reached its full potential. The lack of optimized workflows and isotopically labeled standards limits the acceptance of glycoproteomics quantification. In this paper, we introduce an efficient and streamlined chemoenzymatic synthesis of a library of isotopically labeled glycopeptides of IgG1 which we use for quantification in an energy optimized LC-MS/MS-PRM workflow. Incorporation of the stable isotope labeled N-acetylglucosamine enables an efficient monitoring of all major fragment ions of the glycopeptides generated under the soft collision induced dissociation (CID) conditions which reduces the CVs of the quantification to 0.7-2.8%. Our results document, for the first time, that the workflow using a combination of stable isotope labeled standards with intra-scan normalization enables quantification of the glycopeptides by an electron transfer dissociation (ETD) workflow as well as the CID workflow with the highest sensitivity compared to traditional workflows., This was exemplified by a rapid quantification (13-minute) of IgG1 Fc glycoforms from COVID-19 patients. Graphic Abstract O_FIG O_LINKSMALLFIG WIDTH=200 HEIGHT=115 SRC="FIGDIR/small/501850v1_ufig1.gif" ALT="Figure 1"> View larger version (23K): org.highwire.dtl.DTLVardef@b5b18corg.highwire.dtl.DTLVardef@9b87b9org.highwire.dtl.DTLVardef@eba7ecorg.highwire.dtl.DTLVardef@15da596_HPS_FORMAT_FIGEXP M_FIG C_FIG

biochemistry↗

bioRxiv2020

Ipomoeassin-F inhibits the in vitro biogenesis of the SARS-CoV-2 spike protein and its host cell membrane receptor

In order to produce proteins essential for their propagation, many pathogenic human viruses, including SARS-CoV-2 the causative agent of COVID-19 respiratory disease, commandeer host biosynthetic machineries and mechanisms. Three major structural proteins, the spike, envelope and membrane proteins, are amongst several SARS-CoV-2 components synthesised at the endoplasmic reticulum (ER) of infected human cells prior to the assembly of new viral particles. Hence, the inhibition of membrane protein synthesis at the ER is an attractive strategy for reducing the pathogenicity of SARS-CoV-2 and other obligate viral pathogens. Using an in vitro system, we demonstrate that the small molecule inhibitor ipomoeassin F (Ipom-F) potently blocks the Sec61-mediated ER membrane translocation/insertion of three therapeutic protein targets for SARS-CoV-2 infection; the viral spike and ORF8 proteins together with angiotensin-converting enzyme 2, the host cell plasma membrane receptor. Our findings highlight the potential for using ER protein translocation inhibitors such as Ipom-F as host-targeting, broad-spectrum, antiviral agents.

biochemistry↗