Electrospray ionization mass spectra of acyl carrier protein are insensitive to its solution phase conformation

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• 作者：Peter W. Murphy (1)
  Elden E. Rowland (1)
  David M. Byers (1)
  
• 刊名：Journal of The American Society for Mass Spectrometry
• 出版年：2007
• 出版时间：August 2007
• 年：2007
• 卷：18
• 期：8
• 页码：1525-1532
• 全文大小：894KB
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• 作者单位：Peter W. Murphy (1)
  Elden E. Rowland (1)
  David M. Byers (1)
  
  1. Atlantic Research Centre, Department of Pediatrics, and Departments of Biochemistry and Molecular Biology, Dalhousie University, Halifax, Nova Scotia, Canada
  

文摘

Electrospray ionization mass spectrometry (ESI-MS) can be used to monitor conformational changes of proteins in solution based on the charge state distribution (CSD) of the corresponding gas-phase ions, although relatively few studies of acidic proteins have been reported. Here, we have compared the CSD and solution structure of recombinant Vibrio harveyi acyl carrier protein (rACP), a small acidic protein whose secondary and tertiary structure can be manipulated by pH, fatty acylation, and site-directed mutagenesis. Circular dichroism and intrinsic fluorescence demonstrated that apo-rACP adopts a folded helical conformation in aqueous solution below pH 6 or in 50% acetonitrile/0.1% formic acid, but is unfolded at neutral and basic pH values. A rACP mutant, in which seven conserved acidic residues were replaced with their corresponding neutral amides, was folded over the entire pH range of 5 to 9. However, under the same solvent conditions, both wild type and mutant ACPs exhibited similar CSDs (6+-+ species) at all pH values. Covalent attachment of myristic acid to the phosphopantetheine prosthetic group of rACP, which is known to stabilize a folded conformation in solution, also had little influence on its CSD in either positive or negative ion modes. Overall, our results are consistent with ACP as a “natively unfolded-protein in a dynamic conformational equilibrium, which allows access to (de)protonation events during the electrospray process.