<scp>MS</scp>‐Based Hydroxyl Radical Footprinting: Methodology and Application of Fast Photochemical Oxidation of Proteins (<scp>FPOP</scp>)

Niu, Ben; Gross, Michael L.

doi:10.1002/9781118970195.ch15

Cited by 16 publications

(19 citation statements)

References 293 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…It is important to realize, however, that not all structure‐based MS methods are based solely on the native MS data. The structural information obtained by hydrogen/deuterium exchange (Masson et al, 2019), hydroxyl radical footprinting (Kiselar & Chance, 2018; Niu & Gross, 2019), and covalent labeling (Rajabi, Ashcroft, & Radford, 2015) samples the solution‐phase ion structure. Gross ( Mass spectrometry‐based structural proteomics solves problems in biochemistry and bio‐therapeutics .…”

Section: Introductionmentioning

confidence: 99%

The Ims Paradox: A Perspective on Structural Ion Mobility‐mass Spectrometry

McCabe

Hebert

Shirzadeh

et al. 2020

Mass Spectrometry Reviews

View full text Add to dashboard Cite

Studies of large proteins, protein complexes, and membrane protein complexes pose new challenges, most notably the need for increased ion mobility (IM) and mass spectrometry (MS) resolution. This review covers evolutionary developments in IM‐MS in the authors' and key collaborators' laboratories with specific focus on developments that enhance the utility of IM‐MS for structural analysis. IM‐MS measurements are performed on gas phase ions, thus “structural IM‐MS” appears paradoxical—do gas phase ions retain their solution phase structure? There is growing evidence to support the notion that solution phase structure(s) can be retained by the gas phase ions. It should not go unnoticed that we use “structures” in this statement because an important feature of IM‐MS is the ability to deal with conformationally heterogeneous systems, thus providing a direct measure of conformational entropy. The extension of this work to large proteins and protein complexes has motivated our development of Fourier‐transform IM‐MS instruments, a strategy first described by Hill and coworkers in 1985 (Anal Chem, 1985, 57, pp. 402–406) that has proved to be a game‐changer in our quest to merge drift tube (DT) and ion mobility and the high mass resolution orbitrap MS instruments. DT‐IMS is the only method that allows first‐principles determinations of rotationally averaged collision cross sections (CSS), which is essential for studies of biomolecules where the conformational diversities of the molecule precludes the use of CCS calibration approaches. The Fourier transform‐IM‐orbitrap instrument described here also incorporates the full suite of native MS/IM‐MS capabilities that are currently employed in the most advanced native MS/IM‐MS instruments. © 2020 John Wiley & Sons Ltd. Mass Spec Rev

show abstract

Section: Introductionmentioning

confidence: 99%

The Ims Paradox: A Perspective on Structural Ion Mobility‐mass Spectrometry

McCabe

Hebert

Shirzadeh

et al. 2020

Mass Spectrometry Reviews

View full text Add to dashboard Cite

show abstract

“…Lifetime.-To insure that the labeling reactions do not induce conformational changes during the labeling reaction 49,50 , we examined the correlation between the extent of FPOP modifications and the lifetime (dosage) of the hydroxyl radicals. The lifetime of the radicals can be controlled by varying the concentration of an amino acid radical scavenger 45,51 .…”

Section: Footprinting As Function Of Radicalmentioning

confidence: 99%

Trifluoroethanol Partially Unfolds G93A SOD1 Leading to Protein Aggregation: A Study by Native Mass Spectrometry and FPOP Protein Footprinting

et al. 2020

Self Cite

View full text Add to dashboard Cite

Misfolding of Cu, Zn superoxide dismutase (SOD1) variants may lead to protein aggregation and ultimately amyotrophic lateral sclerosis (ALS). The mechanism and protein conformational changes during this process are complex and remain unclear. To study SOD1 variants aggregation at the molecular level and in solution, we chemically induced aggregation of a mutant variant (G93A SOD1) with trifluoroethanol (TFE) and used both native mass spectrometry (MS) to analyze the intact protein and Fast Photochemical Oxidation of Proteins (FPOP) to characterize the structural changes induced by TFE. We found partially unfolded G93A SOD1 monomers prior to oligomerization and identified regions of the N-terminus, C-terminus, and strands β5, β6 accountable for the partial unfolding. We propose that exposure of hydrophobic interfaces of these unstructured regions serves as a precursor to aggregation. Our results provide a possible mechanism and molecular basis for ALS-linked SOD1 misfolding and aggregation.

show abstract

“…A variety of biophysical techniques have been employed to study protein-protein and protein--ligand interactions (Limpikirati et al, 2018;Liu et al, 2020;Niu & Gross, 2019;Van et al, 2020). Zhang et al used HDX liquid chromatography-mass spectrometry (LC/MS) in addition to cross-linking experiments to identify the binding interface for Nivolumab and its antigen PD-1 (M. M. Zhang et al, 2020).…”

Section: Introductionmentioning

confidence: 99%

Systematic workflow for studying domain contributions of bispecific antibodies to selectivity in multimodal chromatography

Parasnavis

Niu

Aspelund

et al. 2021

Biotech & Bioengineering

Self Cite

View full text Add to dashboard Cite

In this article, a systematic workflow was formulated and implemented to understand selectivity differences and preferred binding patches for bispecific monoclonal antibodies (mAbs) and their parental mAbs on three multimodal cation exchange resin systems. This workflow incorporates chromatographic screening of the parent mAbs and their fragments at various pH followed by surface property mapping and protein footprinting using covalent labeling followed by liquid chromatography-mass spectrometry analysis. The chromatography screens on multimodal resins with the intact mAbs indicated enhanced selectivity as compared to single-mode interaction systems. While the bispecific antibody (bsAb) eluted between the two parental mAbs on most of the resins, the retention of the bispecific transitioned from co-eluting with one parental mAb to the other parental mAb on Capto MMC. To investigate the contribution of different domains, mAb fragments were evaluated and the results indicated that the interactions were likely dominated by the Fab domain at higher pH. Protein surface property maps were then employed to hypothesize the potential preferred binding patches in the solvent-exposed regions of the parental Fabs. Finally, protein footprinting was carried out with the parental mAbs and the bsAb in the bound and unbound states at pH 7.5 to identify the preferred binding patches. Results with the intact mAb analysis supported the hypothesis that interactions with the resins were primarily driven by the residues in the Fab fragments and not the Fc. Furthermore, peptide mapping data indicated that the light chain may be playing a more important role in the higher binding of Parent A as compared with Parent B in these resin systems. Finally, results with the bsAb indicated that both halves of the molecule contributed to binding with the resins, albeit with subtle differences as compared to the parental mAbs. The workflow presented in this paper lays the foundation to systematically study the chromatographic selectivity of large multidomain molecules which can provide insights into improved biomanufacturability and expedited downstream bioprocess development.

show abstract

MS‐Based Hydroxyl Radical Footprinting: Methodology and Application of Fast Photochemical Oxidation of Proteins (FPOP)

Cited by 16 publications

References 293 publications

The Ims Paradox: A Perspective on Structural Ion Mobility‐mass Spectrometry

The Ims Paradox: A Perspective on Structural Ion Mobility‐mass Spectrometry

Trifluoroethanol Partially Unfolds G93A SOD1 Leading to Protein Aggregation: A Study by Native Mass Spectrometry and FPOP Protein Footprinting

Systematic workflow for studying domain contributions of bispecific antibodies to selectivity in multimodal chromatography

Contact Info

Product

Resources

About