From nano- to milimolar: dimerization dissociation constant determination
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| Year of publication | 2022 |
| Type | Conference abstract |
| Citation | |
| Description | The self-association of proteins is the cornerstone of protein regulation, aiding proper functionality and interactome [1]. To properly understand the role of dimerization of protein, only the detection of such dimer is not sufficient, but a quantitative analysis is crucial. A parameter widely used to quantify the self-association is the dissociation constant KD [2]. The KD describes equilibrium between monomers and higher oligomers - in general, the lower the KD, the higher the propensity for oligomerization. However, KD of various proteins differs in several orders of magnitude, providing a challenge in its determination [3]. Here we showcase an array of biophysical methods for coverage of the whole relevant concentration range. For proteins with KD in nM region we optimised Förster resonance energy transfer assay [4]. In µM range an analytical size exclusion combined with multiple angle light scattering can be employed [3]. Finally, we used 19F Trp NMR for evaluation in higher concentrations. Moreover, we show application of such methods on the example of 14–3–3 proteins – cellular regulators connected to oncologic and neurodegenerative diseases [5,6]. |
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