Interactions of nuclear transport factors and surface-conjugated FG nucleoporins: Insights and limitations

Protein-protein interactions are central to biological processes. In vitro methods to examine protein-protein interactions are generally categorized into two classes: in-solution and surface-based methods. Here, using the multivalent interactions between nucleocytoplasmic transport factors and intri...

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Bibliographic Details
Published in:PloS one 2019-06, Vol.14 (6), p.e0217897-e0217897
Main Authors: Hayama, Ryo, Sorci, Mirco, Keating Iv, John J, Hecht, Lee M, Plawsky, Joel L, Belfort, Georges, Chait, Brian T, Rout, Michael P
Format: Article
Language:English
Subjects:
Active Transport, Cell Nucleus
Amino Acid Sequence
Analysis
Antigens
Atomic force microscopy
beta Karyopherins - metabolism
Bioengineering
Biological activity
Biology
Biology and Life Sciences
Biotechnology
Cell Nucleus - metabolism
Desalination
EDTA
Engineering
Enzyme-linked immunosorbent assay
In vitro methods and tests
Interdisciplinary aspects
Laboratories
Ligands
Local transit
Mass transport
Methods
Microbalances
Microscopy
Mutation - genetics
Nuclear Pore Complex Proteins - chemistry
Nuclear Pore Complex Proteins - metabolism
Nuclear transport
Nucleoporins
Physical Sciences
Povidone
Protein Binding
Protein interaction
Protein-protein interactions
Proteins
Quartz crystal microbalance
Quartz Crystal Microbalance Techniques
Quartz crystals
Repetitive Sequences, Nucleic Acid
Research and Analysis Methods
Surface plasmon resonance
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Summary:Protein-protein interactions are central to biological processes. In vitro methods to examine protein-protein interactions are generally categorized into two classes: in-solution and surface-based methods. Here, using the multivalent interactions between nucleocytoplasmic transport factors and intrinsically disordered FG repeat containing nuclear pore complex proteins as a model system, we examined the utility of three surface-based methods: atomic force microscopy, quartz crystal microbalance with dissipation, and surface plasmon resonance. Although results were comparable to those of previous reports, the apparent effect of mass transport limitations was demonstrated. Additional experiments with a loss-of-interaction FG repeat mutant variant demonstrated that the binding events that take place on surfaces can be unexpectedly complex, suggesting particular care must be exercised in interpretation of such data.
ISSN:1932-6203
1932-6203
DOI:10.1371/journal.pone.0217897