DISSECTION OF MULTI-PROTEIN COMPLEXES USING MASS-SPECTROMETRY - SUBUNIT INTERACTIONS IN TRANSTHYRETIN AND RETINOL-BINDING PROTEIN COMPLEXES

Complexes formed between transthyretin and retinol-binding protein pre vent loss of retinol from the body through glomerular filtration. The interactions between these proteins have been examined by electrospray ionization combined with time-of-flight mass analysis. Conditions wer e found whereby complexes of these proteins, containing from four to s ix protein molecules with up to two ligands, are preserved in the gas phase, Analysis of the mass spectra of these multimeric species gives the overall stoichiometry of the protein subunits and provides estimat es for solution dissociation constants of 1.9 +/- 1.0 x 10(-7) M for t he first and 3.5 +/- 1.0 x 10(-5) M for the second retinol-binding pro tein molecule bound to a transthyretin tetramer, Dissociation of these protein assemblies within the gas phase of the mass spectrometer show s that each retinol-binding protein molecule interacts with three tran sthyretin molecules. Mass spectral analysis illustrates not only a cor relation with solution behavior and crystallographic data of a closely related protein complex but also exemplifies a general method for ana lysis of multi-protein assemblies. Proteins Suppl. 2:3-11, 1998. (C) 1 998 Wiley-Liss, Inc.