23 Jul Pichia pastoris-expressed bovine lactoferrin-derived antibacterial fragment BLfA and preparation method thereof
Abstract
The invention relates to preparation methods for anti-bacterial proteins in genetic engineering and particularly relates to a eukaryotic expression protein of a bovine lactoferrin-derived antibacterial fragment BLfA and a preparation method thereof. The encoding genes of the BLfA are cloned from mammary tissue of a Holstein, further, an expression vector is constructed and recombined, so as to realize the expression of the BLfA in Pichia pastoris; the maximum expression level of the BLfA in a shake flask and a 5L fermentation tank is respectively 485 mg/L and 499 mg/L; by using Vitreoscilla hemoglobin (VHb) for co-expressing with rBLfA in a host bacteria cell, the expression level is increased by 23% and reaches 613 mg/L; the rBLfA is subjected to affinity purification, the purity is 93.8%; proved by iron-binding experiments, the Fe<3+> binding capacity of the rBLfA is 0.330, the pH value range for the rBLfA to release Fe<3+> is 7.0-3.5; the rBLfA has a function for inhibiting the growth of staphylococcus aureus, and the MIC (Minimum Inhibitory Concentration) is 6.5 micro mol/L.
Year: 2012
Country: CN
Doc No: 102408480