Enzyme solid-state support assays: a surface plasmon resonance and mass spectrometry coupled study of immobilized insulin degrading enzyme
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Solid-support based assays offer several advantages that are not normally available in solution. Enzymes that are anchored on gold surfaces can interact with several different molecules, opening the way to high throughput array format based assays. In this scenario, surface plasmon resonance (SPR) and mass spectrometry (MS) investigations have often been applied to analyze the interaction between immobilized enzyme and its substrate molecules in a tag-free environment. Here, we propose a SPR-MS combined experimental approach aimed at studying insulin degrading enzyme (IDE) immobilized onto gold surfaces and its ability to interact with insulin. The latter is delivered by a microfluidic system to the IDE functionalized surface and the activity of the immobilized enzyme is verified by atmospheric pressure/matrix assisted laser desorption ionization (AP/MALDI) MS analysis. The SPR experiments allow the calculation of the kinetic constants involved for the interaction between immobilized IDE and insulin molecules and evidence of IDE conformational change upon insulin binding is also obtained.
KeywordsSolid-state assay Surface plasmon resonance Mass spectrometry Insulin degrading enzyme Conformational change
We thank MIUR (FIRB RBNE03PX83, RBIN04L28Y) and “EURAMY: Systemic Amyloidoses in Europe”, 037525 (LSHM-CT-2006-037525) for partial financial support.
- Blomqvist ME-L, Silburn PA, Buchanan DD, Andreasen N, Blennow K, Pedersen NL, Brookes AJ, Mellick GD, Prince JA (2004) Sequence variation in the proximity of IDE may impact age at onset of both Parkinson disease and Alzheimer disease. Neurogenetics 5:115–119. doi: 10.1007/s10048-004-0173-4 PubMedCrossRefGoogle Scholar
- Darnell JE, Lodish H, Baltimore D (1990) Molecular cell biology. Scientific American Books, New YorkGoogle Scholar
- Drobny GP, Long JR, Karlsson T, Shaw W, Popham J, Oyler N, Bower P, Stringer JD, Mehta GM, Stayton PS (2003) Structural studies of biomaterials using double-quantum solid-state NMR spectroscopy. Annu Rev Phys Chem 54:531–571. doi: 10.1146/annurev.physchem.54.011002.103903 PubMedCrossRefGoogle Scholar
- Farris W, Mansourian S, Chang Y, Lindsley L, Eckman EA, Frosch MP, Eckman CB, Tanzi RE, Selkoe DJ, Guénette S (2003) Insulin-degrading enzyme regulates the levels of insulin, amyloid β-protein, and the β-amyloid precursor protein intracellular domain in vivo. Proc Natl Acad Sci USA 100:4162–4167. doi: 10.1073/pnas.0230450100 PubMedCrossRefGoogle Scholar
- Grasso G, D’Agata R, Rizzarelli E, Spoto G, D’Andrea L, Pedone C, Picardi A, Romanelli A, Fragai M, Yeo KJ (2005) Activity of anchored human matrix metalloproteinase-1 catalytic domain on Au (111) surfaces monitored by ESI-MS. J Mass Spectrom 40:1565–1571. doi: 10.1002/jms.929 PubMedCrossRefGoogle Scholar
- Homola J (ed) (2006) Surface plasmon resonance based sensors. Springer, BerlinGoogle Scholar
- Im H, Manolopoulou M, Malito E, Shen Y, Zhao J, Neant-Fery M, Sun C-Y, Meredith SC, Sisodia SS, Leissring M, Tang W-J (2007) Structure of substrate-free human insulin-degrading enzyme (IDE) and biophysical analysis of ATP-induced conformational switch of IDE. J Biol Chem 282:25453–25463. doi: 10.1074/jbc.M701590200 PubMedCrossRefGoogle Scholar
- Lahiri J, Isaacs L, Tien J, Whitesides GM (1999) A strategy for the generation of surfaces presenting ligands for studies of binding based on an active ester as a common reactive intermediate: a surface plasmon resonance study. Anal Chem 71:777–790. doi: 10.1021/ac980959t PubMedCrossRefGoogle Scholar
- Sharff AJ, Rodseth LE, Spurlino JC, Quiocho FA (1992) Crystallographic evidence of a large ligand-induced hinge-twist motion between the two domains of the maltodextrin binding protein involved in active transport and chemotaxis. Biochemistry 31:10657–10663. doi: 10.1021/bi00159a003 PubMedCrossRefGoogle Scholar