Utility of phenylhydrazine (PHN) labeling for structural studies of fucosylated N-glycans by tandem MALDI mass spectrometry (MS) in the positive ion mode is proposed. PHN-tag influences the production of specific ion types, and the MS/MS fragmentation pattern provides useful structural information. 

Zbyněk Zdráhal Research Group

Significance

Fucosylation is a common modification, and its site in glycans refers to different normal and pathological processes. Despite intensive research, there is still a lack of methods to discriminate unambiguously the fucose position in one-step. In this work, we propose utility of phenylhydrazine (PHN) labeling for structural studies of fucosylated N-glycans by tandem MALDI mass spectrometry (MS) in the positive ion mode. PHN-tag influences the production of specific ion types, and the MS/MS fragmentation pattern provides useful structural information. All types of core fucosylated N-glycans have produced two abundant ions consistent with B- and C-glycosidic cleavages corresponding to the loss of the FucGlcNAcPHN residue with a mass 457 and 441 Da from the parent ions. These types of fragment ions in N-glycans without a core fucose were associated with the loss of the GlcNAcPHN unit (311 and 295 Da), and fucose cleavage followed the loss of the chitobiose residue. Since diagnostic useful cleavages produce peaks with significant intensities, this approach is also beneficial for rapid recognition of antenna from core fucosylation in glycans detected with low abundances. The practical applicability of the approach is demonstrated on the analysis of multifucosylated N-glycans detected with lower abundances in lung cancer samples

Lattová, E., Skřičková, J., and Zdráhal, Z.: Applicability of Phenylhydrazine Labeling for Structural Studies of Fucosylated N-Glycans, Anal. Chem. (2019) 91(13),7985-7990. https://doi.org/10.1021/acs.analchem.9b01321

Potato virus Y (PVY) belongs to the most economically important pathogens. The collaborative research project of the National Institute of Chemistry (Ljublana, Slovenia) and Cryo-Electron Microscopy Core Facility at CEITEC MU reveals the structure of the PVY coat protein (CP) and the PVY virus like particle at near-atomic resolution. The data show a novel luminal interplay between the extended carboxy-terminal CP regions in the virion and describe RNA-CP interactions important for helical conformation and stability of the virus.

Marjetka Podobnik Research Group

Significance

PVY is ranked as fifth in the top 10 most economically important plant viruses and is the most important viral pathogen of potato worldwide. The virus causes potato tuber necrotic ringspot disease, which can result in up to 70% yield reduction, and severely affects other economically important solanaceous crops. Despite extensive availability of data on PVY’s genome and pathogenicity, there has been no high-resolution structuralinformation for this virus. Because of the extreme economic importance of PVY, and the urgent need for structural data to better understand mechanisms of viral infectivity, we have examined in detail the structure of the PVY virion and its CP. We have determined the high-resolution electron cryo-microscopy structures of the PVY virion and a recombinant PVY-based RNA-free virus-like particle (VLP). This provides a new and detailed insight into the RNA-supported helical viral capsid architecture featuring an extended C-terminal region of CP, which is tightly packed in a unique fashion in the virion lumen. In addition, using extensive biochemical, biophysical, and computational characterization, as well as structure-­based mutagenesis, we identified regions of CP that affect VLP filament assembly. Moreover, the biological activities of the CP’s N- and C-terminal regions for virus infectivity were explored by measuring the accumulation of viral RNA and systemic movement of selected PVY mutants in plants.

Kežar, A., Kavčič, L.,  Polák, M., Nováček, J., Gutiérrez-Aguirre, I., Tušek Žnidarič, M., Coll, A., Stare, K., Gruden, K., Ravnikar, M., Pahovnik, D., Žagar, E., Merzel, F., Anderluh,G., and Podobnik, M.: Structural basis for the multitasking nature of the potato virus Y coat protein, Sci. Adv. (2019) 5(7), eaaw3808, DOI: 10.1126/sciadv.aaw3808