Pharmaceutical Sciences Faculty Publications

Low Temperature Aqueous Electrospray Ionization Mass Spectrometry of Noncovalent Complexes

Document Type

Article

Publication Date

6-1-1998

Journal Title

Journal of the American Society for Mass Spectrometry

ISSN

1044-0305

Volume

9

Issue

6

First Page

580

Last Page

584

DOI

10.1016/S1044-0305(98)00019-1

PubMed ID

9879371

Abstract

In the present study we describe conditions that permit the characterization of noncovalent protein-substrate complexes in aqueous solution by microspray electrospray ionization-mass spectrometry (ESI-MS), using a heated transfer capillary at low temperature (45 degrees C). Specifically, we examined the binding of calmodulin to two polypeptides; the calmodulin-binding domain of calmodulin-dependent protein kinase II (CamK-II) and melittin. Calmodulin, a well known calcium-binding protein, binds to a number of small amphipathic peptides in a calcium-dependent manner. Our results directly show that both peptides form equimolar complexes with calmodulin only in the presence of calcium. The stoichiometry necessary for the formation of each complex was 1:1:4 for calmodulin:peptide (melittin or CamK-II):Ca2+, respectively. Furthermore, it is demonstrated that the detection of the complex in ESI-MS is source temperature dependent.

Keywords

Calcium-calmodulin-dependent protein kinases, calmodulin, mass spectrometry, melitten, temperature

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