Mass Spectrometry of Biopolymers
CS

Service

Routine proteomic analyses

Our group provide service proteomic analyses based on mass spectrometry detection. We offer following proteomic analyses:
  • Intact mass analysis
            molecular weight determination of protein or oligonucleotides by MALDI-TOF or ESI-TOF
  • Protein identification
            including protein complexes characterization from gel or solution
  • Characterization of posttranslational modifications
            phosphorylation, acetylation, methylation etc.
  • Protein quantification
            relative quantification (label-free, SILAC labeling)

Specialized service analyses

In our group we aslo implement novel structural proteomics approaches.

  • native MS
  • protein covalent labelling
  • protein crosslinking
  • hydrogen-deuterium exchange MS
These complementary methods allow to study protein structures and characterize not only protein-protein interactions but also interactions of proteins with other biological molecules.

Ion mobility spectrometry (IMS) of amyloid beta peptide (1-42). Individual chrge states of monomers and oligomers are labeled.
Phosphorylase B labeled by D2O in two time points (20 seconds and 2 hours). Obtained data were shown in phosphorylase B structure where the relative deuteration uptake of identified regions was expressed in a color scale.

Projects

Our group is involved in numerous projects where the MS-based proteomic approaches is required. These projects deal with, for example, the research of Hepatitis B viral infection or analysis of rhomboid dependent proteome changes, etc.

A) Total ion current chromatogram obtained for the sample of mouse lysate. The MS/MS scan of molecule with retention time of 48.81 min is shown in red rectangle. B) Volcano plots as a result of student's t-test comparison of shotgun LC-MS/MS analysis of proteins co-immunoprecipitated with HBe-HA (left) and HBe3CA-HA (right). Published in Zábranská et al., The FEBS Journal, 2021.

In our research projects we focus on:

  •     analysis of lipoproteins/lipopeptides and membrane proteins
Separation and identification of lipoproteins/lipopeptides is rather challenging due to the limited solubility of lipo-modified segments in water-based solutions, whose are exclusively used within standard proteomic workflows. Therefore, we focus on the implementation of new workflows to the increase the yield of lipo-modified and hydrophobic segments of the proteins.

The fragmentation spectrum of the myristoylated peptide GVSGSK obtained by CID and HCD fragmentation technique. In the case of HCD both indicator fragments of myristoylation b0 (m/z 211.205) and b1 (m/z 268.227) were detected.

  •      Application of the methods of structural proteomics on the field of retroviral life cycle studies 

Although the life cycle of retroviruses is intensively studied worldwide, some of the mechanism used by retroviruses in host cells are still not elucidated. This refers to the interaction of newly formed virus particles with cell membrane before they are released from the host cell. To clarify the mechanism that retroviruses use in this step, we apply structural proteomics methods, mainly protein cross-linking and the hydrogen-deuterium exchange method.