Abstract
Seed proteins have been evaluated for their nutritive value and biological activity. Proteases and proteinase inhibitors have been of immense benefit to both agriculture and therapeutic purposes. The seed proteins of Blighia sapida was evaluated for both structure and biological activity in this study. The matured dried seeds were pulverized and sequentially extracted using 10 mM Tris/HCl pH 7.4, 10 mM ammonium acetate and 10 mM sulphuric acid. Crude protein extracts were concentrated and the protein concentrations were estimated. Proteins were purified by 70% ammonium sulphate precipitation, Sephadex G50 reversed phase chromatography and finally by HPLC on a C18 column. Two bands were obtained from SDS-PAGE electrophoresis and they were identified by ESI/MS using in gel tryptic digestion. The seed protein from B. Sapida consists of two single polypeptide chains each with mass of about 24 to 27 KDa as established by a combination of SDS-PAGE and ESI/MS. Proteins exhibited protease activity, which was confirmed by zymography. Protease activity was characterized for effect of temperature, pH, divalent metal ions and chelating agents. The crude protein from the seed of B. sapida showed antimicrobial activity and the antifungal activity was comparable with the reference drug, Ticonazole. Keywords: Blighia sapida, chromatography, protease activity, in-gel trypsin digestion/mass spectrophotometry, antimicrobial activity. African Journal of Biotechnology , Vol 13(29) 2996-3007
Highlights
Ackee, the national fruit of Jamaica, is a food staple in many Jamaican diets (Sharma et al, 2009)
MO, USA); ammonium per sulfate (APS); bromophenol blue (3,3,5,5-tetrabromophenolsulfonephthalein); tetramethylethylene diamine (TEMED); 2-mercaptoethanol; glycerol, sodiumdodecyl sulphate (SDS), DL-dithiotreitol were purchased from Sigma as well
Obtained chromatograms from size exclusion chromatography are shown as (Figures Ia, b and c) while the protein concentrations of the concentrated pulled chromatographic fractions are shown on Table 1
Summary
The national fruit of Jamaica, is a food staple in many Jamaican diets (Sharma et al, 2009). Blighia sapida is a woody perennial multipurpose fruit tree species native to the Guinean forests of West Africa. 4.0 International License splits open while still on the tree to reveal 3glassy black seeds surrounded by a thick, oily, yellow aril. An association between ackee poisoning and Jamaican vomiting sickness was made in 1875 with a water-soluble toxic material in the seed and pods of the ackee fruit (Joskow et al, 2006; Moya, 2001; Gaillard et al, 2011)
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