Tinh sạch phytase của vi khuẩn được phân lập và tuyển chọn từ dạ cỏ cừu
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Abstract
The research was conducted to isolate, and select bacteria from sheep rumen which were capable of producing and purifying phytase including methods of protein precipitation by ammonium sulfate salt 80% and ion exchange chromatography on DEAE-cellulose gel. The molecular weight of the protein was assessed by SDS-PAGE electrophoresis. Among 16 isolates of bacteria from sheep rumen which were capable for degrading phytate, the strain C8 was determined to create the highest protein and phosphate content of 0.320 mg/mL, 11.76 mg/mL, respectively. The nucleotide sequence of 16S rDNA region between the bacterial strain C8 isolated from the sheep rumen and others on the NCBI genbank were compared. The results showed that the C8 bacterial strain was similar 93.16% with the Bacillus rugosus strain SPB7. Phytase fraction F1 was obtained with recovery efficiency of 13.2%, purity 3.04 times, total activity of 8,733 U, specific activity of 280 U/mg protein and molecular weight 34.9 kDa.
Tóm tắt
Nghiên cứu được thực hiện để phân lập, tuyển chọn vi khuẩn từ dạ cỏ cừu có khả năng tổng hợp phytase và tinh sạch phytase bao gồm phương pháp kết tủa protein bằng muối ammonium sulphate 80% và kết hợp sắc ký trao đổi ion trên gel DEAE-cellulose. Khối lượng phân tử protein được đánh giá bằng phương pháp điện di SDS-PAGE. Trong 16 chủng vi khuẩn dạ cỏ cừu có khả năng phân giải phytate, dòng vi khuẩn C8 được xác định có hàm lượng protein và phosphate cao nhất lần lượt là 0,320 mg/mL, 11,76 mg/mL. Kết quả giải trình tự vùng 16S rDNA của dòng vi khuẩn C8 khi so sánh với ngân hàng gen NCBI và đặc điểm vi khuẩn phân lập được từ dạ cỏ cừu cho thấy dòng vi khuẩn C8 xác định được mức tương đồng với dòng vi khuẩn Bacillus rugosus strain SPB7 ở mức tương đồng 93,16%. Phân đoạn phytase F1 thu được đạt hiệu suất thu hồi là 13,2 %, độ tinh sạch gấp 3,04 lần, hoạt tính tổng 8.733 U, hoạt tính đặc hiệu là 280 U/mg protein và khối lượng phân tử 34,9 kDa.
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