An efficient method for purification of nonspecific lipid transfer protein‐1 from rice seeds using kiwifruit actinidin proteolysis and ion exchange chromatography

Plant nonspecific lipid transfer proteins are small basic proteins that transport phospholipids between membranes and are subdivided into two subfamilies, nsLTP₁ (9 kDa) and nsLTP₂ (7 kDa). LTPs have potential application in the defense reactions against pathogens and the drug delivery systems. Many efforts have been made for purification of different nsLTPs from various plants; however, most of them used successive purification procedures. We have developed a relatively simple and efficient method for the purification of rice nsLTP₁, based on the proteolytic activity of kiwifruit actinidin on the rice seed extract and one‐step chromatographic procedure on a CM‐Sepharose column. The purity of protein was determined by reversed‐phase high‐performance liquid chromatography (RP‐HPLC) and sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS‐PAGE). The isolated LTP₁ migrated as a homogenous polypeptide with molecular mass of 9 kDa that confirms the efficiency of actinidin on the digestion of major contaminations present in the rice seed extract without any harmful effect on the LTP₁. The advantages of using proteolytic activity of actinidin in purifying rice LTP₁ includes the reduced separation time allowing the purification of LTP₁ in one‐step chromatographic procedure, low costing, high efficiency, and the relative simplicity of the method.