Methodology for measuring conformation of solvent-disrupted protein subunits using T-WAVE ion mobility MS: An investigation into eukaryotic initiation factors |
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Authors: | Julie A Leary Matthew R Schenauer Raluca Stefanescu Armann Andaya Brandon T Ruotolo Carol V Robinson Konstantinos Thalassinos James H Scrivens Masaaki Sokabe John W B Hershey |
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Institution: | aDepartment of Molecular and Cellular Biology, University of California at Davis, Davis, California, USA;bDepartment of Chemistry, University of Cambridge, Cambridge, United Kingdom;cDepartment of Biological Sciences, University of Warwick, Coventry, United Kingdom;dDepartment of Biochemistry and Molecular Medicine, School of Medicine, University of California at Davis, Davis, California, USA |
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Abstract: | The methodology developed in the research presented herein makes use of chaotropic solvents to gently dissociate subunits
from an intact macromolecular complex and subsequently allows for the measurement of collision cross section (CCS) for both
the recombinant (R-eIF3k) and solvent dissociated form of the subunit (S-eIF3k). In this particular case, the k subunit from
the eukaryotic initiation factor 3 (eIF3) was investigated in detail. Experimental and theoretical CCS values show both the
recombinant and solvent disrupted forms of the protein to be essentially the same. The ultimate goal of the project is to
structurally characterize all the binding partners of eIF3, determine which subunits interact directly, and investigate how
subunits may change conformation when they form complexes with other proteins. Research presented herein is the first report
showing retention of solution conformation of a protein as evidenced by CCS measurements of both recombinant and solvent disrupted
versions of the same protein. |
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