Native mass spectrometry (nMS), in which intact protein or nucleoprotein complexes are directly measured, commonly involves exchange of nonvolatile electrolytes (NaCl, PBS, Tris) for volatile electrolytes such as ammonium acetate. This can be time-consuming if offline buffer exchange spin cartridges are used. Furthermore, some samples may not be stable if left in nonvolatile buffer for more than just a very short period of time. This problem can be avoided if an LC-compatible online buffer exchange column is used to separate protein complex from salt. In this experiment, no attempt is made to perform size exclusion chromatography to separate protein complexes of different sizes but rather any protein complex that is injected onto the system is simply separated from low mass salts that would otherwise product heavily adducted protein complexes that appear as broad peaks in the mass spectrum. This webinar will focus on the development of online buffer exchange (OBE) for nMS applications and will describe how the experiment is implemented and how it can be extended by coupling to affinity separation (e.g., IMAC-OBE) to be used to optimize protein overexpression.

About the presenter

Vicki Wysocki, Ph.D., Professor, Dept of Chemistry and Biochemistry, The Ohio State University

Vicki Wysocki, Ph.D., Professor, Dept of Chemistry and Biochemistry,
The Ohio State University

Dr. Vicki Wysocki received her BS in Chemistry from Western Kentucky University in 1982 and her Ph.D. in Chemistry from Purdue University in 1987. After pursuing postdoctoral studies at Purdue and at the Naval Research Laboratory as a National Research Council Fellow, she joined Virginia Commonwealth University as an Assistant Professor in 1990. She was promoted to Associate Professor in 1994. Vicki joined the University of Arizona in 1996 and was promoted to Professor in 2000. Most recently she was Chair of the Department of Chemistry and Biochemistry at Arizona. Vicki joined Ohio State in August 2012 as an Ohio Eminent Scholar and Director of the OSU Campus Chemical Instrument Center.

Key learning objectives

  • The use of online buffer exchange for high throughput native mass spectrometry
  • Online buffer exchange nMS allows to directly screen structural features of large biomolecules even if such biomolecules are not very stable in MS friendly buffers
  • How the coupling of affinity separation (e.g., IMAC-OBE) to online-buffer exchange can be used to optimize protein overexpression

Who should attend

  • MS practitioners
  • Researchers/R&D managers
  • Laboratory managers/directors/supervisors
  • Laboratory technicians/operators
  • Structural biologists
  • Scientists in biopharma engaged in biotherapeutic characterization

 
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