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PMID: 15642257 Published · ppublish English Journal Article Research Support, Non-U.S. Gov't

When monomers are preferred: a strategy for the identification and disruption of weakly oligomerized proteins.

Structure (London, England : 1993) ·Vol. 13 ·No. 1 ·2005-01-00 ·Pages 7-15

Tong Y, Hughes D, Placanica L, Buck M

Abstract

Oligomerization is important for the structure and function of many proteins, but frequently complicates their characterization. It is often desirable to obtain the protein in monomeric form. Here, we report a strategy that allows the generation of monomers from weakly associated oligomers but does not require knowledge of the three-dimensional structure of the protein. The dynamics of protein association are used in solution NMR spectroscopy to identify regions of the polypeptide chain that are likely to be responsible for the interaction. Protein sequence analysis further refines the selection, as conserved sites with moderate hydrophobicity are targeted for modification. Gel filtration and activity assays straightforwardly reveal the consequences of the change and are used to screen for the desired mutants. The strategy is demonstrated for the Rac1 binding domain of plexin-B1. A monomeric variant is generated which preserves the Rac1 binding activity and the wild-type protein structure.

MeSH Terms
Amino Acid Sequence Binding Sites Chromatography, Gel Consensus Sequence Dimerization Hydrophobic and Hydrophilic Interactions Molecular Sequence Data Mutagenesis, Site-Directed Nuclear Magnetic Resonance, Biomolecular Point Mutation Protein Binding Protein Structure, Secondary Protein Structure, Tertiary Proteins/chemistry,genetics,metabolism Sequence Analysis, Protein Sequence Homology, Amino Acid Solutions
Chemicals
Proteins Solutions
Authors & Affiliations
4 authors, click to expand affiliations / ORCID
Tong Yufeng
Department of Physiology and Biophysics, Case Western Reserve University School of Medicine, Cleveland, Ohio 44106, USA.
Hughes David
Placanica Lisa
Buck Matthias
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Article Info
Journal
Structure (London, England : 1993)
Abbr.
Structure
ISSN
0969-2126
DOI
10.1016/j.str.2004.10.018
Published
2005-01-00
Pages
7-15
Language
English
Region
United States
NLM ID
101087697
PMCID
PMC2765720
Subset
IM
Grants
NIGMS NIH HHS · R01 GM073071 · United States
NIGMS NIH HHS · R01 GM073071-01 · United States
External Links
PubMed source DOI: 10.1016/j.str.2004.10.018 PMC Full Text View journal details
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