Investigation of intact protein complexes by mass spectrometry

Abstract

I. Introduction 00 II. Electrospray Ionization of Biomacromolecules 00 III. Mass Analyzers for Biomacromolecular Mass Spectrometry 00 A. Collisional Cooling and/or Focusing 00 B. Alternative Mass Analyzers 00 IV. Solution‐Phase Properties of Proteins Complexes 00 A. Protein–Protein Interactions 00 B. Cooperativity and Cofactors 00 C. Dynamics of Assembly 00 V. Gas‐Phase Properties of Protein Complexes 00 A. Tandem Mass Spectrometry 00 B. Charge Separation in Protein Dissociation 00 VI. Future Outlook 00 Acknowledgments 00 References 00 Mass spectrometry has grown in recent years to a well‐accepted and increasingly important complementary technique in structural biology. Especially electrospray ionization mass spectrometry is well suited for the detection of non‐covalent protein complexes and their interactions with DNA, RNA, ligands, and cofactors. Over the last decade, significant advances have been made in the ionization and mass analysis techniques, which makes the investigation of even larger and more heterogeneous intact assemblies feasible. These technological developments have paved the way to study intact non‐covalent protein–protein interactions, assembly and disassembly in real time, subunit exchange, cooperativity effects, and effects of cofactors, allowing us a better understanding of proteins in cellular processes. In this review, we describe some of the latest developments and several highlights. © 2004 Wiley Periodicals, Inc., Mass Spec Rev

Gas phase noncovalent protein complexes that retain solution binding properties: Binding of xylobiose inhibitors to the β-1, 4 exoglucanase from Cellulomonas fimi

“…Peaks from enzyme (E) and complex (EI) are seen. A narrow distribution of charge states from ϩ9 to ϩ12, with ϩ10 being the most intense, is formed as expected°for°the°protein°in°its°native°conformation° [25,30].°The°peaks°of°the°complex°(EI)°are°30°to°40%°as intense as those of the enzyme (E) for the same charge state, even though, from the solution K i , it is calculated that°almost°all°the°enzyme°was°in°the°complexed°form. The gas-phase abundances do not reflect the solution equilibria.…”

Ligand-induced monomerization of Allochromatium vinosum cytochrome c′ studied using native mass spectrometry and fluorescence resonance energy transfer

“…Native MS allowed simultaneous detection of monomeric, dimeric and unfolded forms of cyt-c¢ and was therefore able to monitor the CO-induced monomerization of cyt-c¢. MS also allowed the detection of the CO-bound form of cyt-c¢, which to our knowledge is the first time that the binding of a gaseous ligand to a protein has been observed with MS. An explanation for the partial preservation of the CO-bound complex is that the bound CO molecule is located deep inside the channel that gives access to the distal coordination site [26].…”

“…Recent developments in MS instrumentation have enabled the characterization of native, folded proteins and large protein complexes that are held together by weak noncovalent interactions [25,26]. Several studies have reported the use of this so-called native MS to study protein folding and cofactor binding [26,27].…”

“…Several studies have reported the use of this so-called native MS to study protein folding and cofactor binding [26,27]. Since MS, unlike many spectroscopic techniques, can detect all species present in solution, we explored its suitability for monitoring the COinduced conformational changes in cyt-c¢.…”

Effect of tanshinone IIA on the noncovalent interaction between warfarin and human serum albumin studied by electrospray ionization mass spectrometry

“…Moreover, NMR is limited to molecules with molecular weight not exceeding 40 kDa; furthermore, not all proteins are easy to crystallize for X-ray crystallography detection [26]. Electrospray ionization mass spectrometry (ESI-MS) has recently emerged as an attractive alternative [27][28][29][30][31]. Although unable to provide direct structural data as NMR and X-ray crystallography, ESI-MS does provide important stoichiometry information of the complex formed between two binding partners.…”