Expression and purification of class two small heat shock proteins from maize

CHED 839

Christopher T. Nau, cnau09@wooster.edu1, Virginia B. Pett1, Hannah S. Tims1, and Robert A. Bouchard2. (1) Department of Chemistry, The College of Wooster, 943 College Mall, Wooster, OH 44691, (2) Department of Horticulture and Crop Science, Ohio Agricultural Research & Development Center, 1680 Madison, Wooster, OH 44691
Small heat shock proteins (sHsps) are a classification of chaperones that bind to a substrate in order to reduce undesirable aggregation, ultimately interacting with other ATP-activated chaperones to encourage proper re-folding of denatured proteins. To this end, sHsps are often expressed during increased stress or temperature. The goal of the research was to maximize yield during expression and purification of three small heat shock proteins from Zea maize: 17.0, 17.6, 17.8 (classified according to their size in kilodaltons). Affinity chromatography using an intein tag was used as the first step of purification, while size exclusion chromatography (SEC) was explored as a secondary purification step. SEC proved to be an effective method for separating the target protein. However, the accompanying loss of protein led to further purification attempts using hydroxyapatite column material. Other factors were also analyzed, including detergent concentration, pH of concentration, and DNA digestion enzymes.