© 2021 Institute of Food Science and TechnologySesame seed is mostly utilised for its oil but also the waste of the oil processing; the seed meal has also significant potential to be used as an alternative protein source. In this study, the goal is to produce sesame seed protein by using three different techniques; alkaline, salt and enzyme-assisted extraction. A comprehensive physicochemical characterisation of the extracts was performed. Total and soluble protein contents, emulsification activity & emulsion stability, FTIR spectroscopy, hydration behaviour and gelling ability experiments by TD-NMR were conducted for all extracted proteins. Also, SDS-PAGE experiments were performed to observe the effect of extraction conditions on protein folding. Overall, the aqueous phase of enzyme-assisted extracted proteins (E-ACP) had the highest protein content and solubility, which resulted in other improved physicochemical properties. Salt extracted samples were ‘salted-out’, therefore, had poor physicochemical properties. TD-NMR experiments further confirmed the solubility and gelling ability results by measuring the change in the T2 spin relaxation times. Additionally, FTIR spectroscopy confirmed the most critical peaks for the proteins; Amide I (C=O stretching) and Amide II (N–H bending). In summary, depending on the physicochemical property of interest, different extraction methods yielded proteins with different properties.