Napin from mustard (Brassica juncea L.) is a seed storage protein consisting of two subunits linked through disulfide bonds and is predominantly helical in nature. Resistance to trypsin digestion and allergenicity limit its food applications. The role of disulfide linkages, electrostatic as well as hydrophobic interactions, in napin stability have been investigated through spectroscopic methods, employing different fluorescent probes and additives. The subunits are hydrophilic in nature and possess extended structure. With the addition of 0.5 M NaCl, the surface hydrophobicity of napin decreases, whereas the helical content increases by 25%. In the presence of NaCl, emission maximum shifts toward shorter wavelength and the Stern-Volmer constant decreases from 6.5 to 3.4 M-1, indicating compaction of napin. Na2SO4 has no significant effect on the structure due to the lack of a hydrophobic core. In the presence of monohydric alcohols and trifluoroethanol, there is an increase in ordered structure. These studies indicate that the structure of napin, which is hydrophilic in nature, is stabilized by electrostatic interactions, in addition to disulfide linkages.