Dataset Information

Hydroxyl radical footprinting analysis of a human haptoglobin-hemoglobin complex

ABSTRACT: Methods of structural mass spectrometry have become more popular to study protein structure and dynamics. Among them, fast photochemical oxidation of proteins (FPOP) has several advantages such as irreversibility of modifications and more facile determination of the site of modification with single residue resolution. In the present study, FPOP analysis was applied to study the hemoglobin (Hb) – haptoglobin (Hp) complex allowing identification of respective regions altered upon the complex formation. The footprinting using a timsTOF Pro mass spectrometer allowed to obtain structural information for 84 and 76 residues in Hp and Hb, respectively, including statistically significant differences in the modification extent below 0.3 %. The most affected residues upon the complex formation were Met76 and Tyr140 in Hbα, and Tyr280 and Trp284 in Hpβ. The data allowed determination of amino acids directly involved in Hb – Hp interactions and those located outside of the interaction interface yet affected by the complex formation. Also, previously modeled interaction between Hb βTrp37 and Hp βPhe292 was not confirmed by our data.

INSTRUMENT(S): timsTOF Pro

ORGANISM(S): Homo Sapiens (human)

SUBMITTER: Dmitry Loginov

LAB HEAD: Petr Novak

PROVIDER: PXD021621 | Pride | 2021-12-30

REPOSITORIES: Pride

ACCESS DATA

Publications

Hydroxyl radical footprinting analysis of a human haptoglobin-hemoglobin complex.

Loginov Dmitry S DS Fiala Jan J Brechlin Peter P Kruppa Gary G Novak Petr P

Biochimica et biophysica acta. Proteins and proteomics 20211103 2

Methods of structural mass spectrometry have become more popular to study protein structure and dynamics. Among them, fast photochemical oxidation of proteins (FPOP) has several advantages such as irreversibility of modifications and more facile determination of the site of modification with single residue resolution. In the present study, FPOP analysis was applied to study the hemoglobin (Hb) - haptoglobin (Hp) complex allowing identification of respective regions altered upon the complex forma ...[more]

PMID: 34742912

Similar Datasets

Project description:The present study aimed to develop and optimize cell derived membrane-coated nanostructures by applying a design of experiments (DoE) to improve the therapeutic index by homotypic targeting in cancer cells. The protein composition of the extracted cell membrane from tumoral cells were assessed by mass spectrometry-based proteomics. PLGA-based nanoparticles encapsulating temozolomide (TMZ NPs) were successfully developed. The coating technology applying the isolated U251 cell membrane (MB) was optimized using a fractional two-level three-factor factorial design. All the formulation runs were systematically characterized regarding their diameter, polydispersity index (PDI), and zeta potential (ZP). Experimental conditions generated by DoE were also subjected to morphological studies using negative-staining transmission electron microscopy (TEM). MicroRaman, Fourier-Transform Infrared (FTIR) spectroscopies and Confocal microscopy were used as characterization techniques for evaluating the NP-MB nanostructures. Internalization studies were carried out to evaluate the homotypic targeting ability. The results have shown that nearly 80% of plasma membrane proteins were retained in the cell membrane vesicles after the isolation process, including key proteins to the homotypic binding. DoE analysis considering acquired TEM images reveals that condition run five should be the best-optimized procedure to produce the biomimetic cell-derived membrane-coated nanostructure (NP-MB). Raman, FTIR, and confocal characterization results have shown the successful encapsulation of TMZ drug and provided evidence of the effective coating applying the MB. Cell internalization studies corroborate the proteomic data indicating that the optimized NP-MB achieved specific targeting of homotypic tumor cells. The structure should retain the complex biological functions of U251 natural cell membranes while exhibiting physicochemical properties suitable for effective homotypic recognition. Together, these findings provide coverage and a deeper understanding regarding the dynamics around extracted cell membrane and polymeric nanostructures interactions and an in-depth insight into the cell membrane coating technology and the development of optimized biomimetic and bioinspired nanostructured systems.

Dataset Information

Hydroxyl radical footprinting analysis of a human haptoglobin-hemoglobin complex

Publications

Hydroxyl radical footprinting analysis of a human haptoglobin-hemoglobin complex.

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure