Tối ưu điều kiện biểu hiện nhằm nâng cao khả năng sinh tổng hợp Chitinase chủng vi khuẩn tái tổ hợp bằng phương pháp bề mặt đáp ứng

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Tác giả: Văn Quyền Đồng, Đình Bình Ngô, Thị Thu Hà Trịnh

Ngôn ngữ: vie

Ký hiệu phân loại: 589.95 [Unassigned]

Thông tin xuất bản: Sinh học, 2018

Mô tả vật lý: 115-123

Bộ sưu tập: Metadata

ID: 492067

Thêm vào giỏ

Chitinase (EC 3.2.1.14) thuộc nhóm enzyme thủy phân (hydrolase), là enzyme thủy phân chitin thành chitobiose hay chitotriose qua việc xúc tác sự phân giải liên kết 1,4 glucoside giữa C1 và C4 của hai phân tử N-acetyl glucosamine liên tiếp nhau trong chitin. Nghiên cứu đã xác định điều kiện tối ưu cho lên men sản xuất chitinase tái tổ hợp bao gồm: nồng độ lactose 6,15 mM
thời gian cảm ứng 8,12 giờ và mật độ tế bào tại thời điểm cảm ứng là OD600nm=0,87 với giá trị chitinase đạt 7,49 U/ml. Tiến hành lên men chủng tái tổ hợp theo điều kiện tối ưu, hoạt tính chitinase theo thực tế đạt 7,54 U/ml, cao gấp 1,32 lần so với trước tối ưu (hoạt tính đạt 5,71 U/ml). Ngoài ra, sử dụng chất cảm ứng lactose thay thế ITPG còn giúp giảm giá thành sản phẩm chitinase tái tổ hợp.Chitinases (EC 3.2.1.14) are enzymes that degrade chitin. These enzymes are gaining importance for their wide-ranging applications including the preparation of pharmaceutically important chitooligosaccharides and N-acetyl D glucosamine, preparation of single-cell protein, isolation of protoplasts from fungi and yeast, control of pathogenic fungi, treatment of chitinous waste, mosquito control and morphogenesis and especially in agriculture fields to control pathogens. Previously, we have cloned chiA gene encoding chitinase for overexpresion in E. coli. Herein, we optimized the expression conditions for improvement of chitinase biosynthesis in recombinant E. coli strain using response surface method (RSM) based on Design-Expert 10.1 software (Stat-Ease, Inc., Minneapolis, USA. Fermentation conditions such as inducer concentrate, induction time and cell density at the time of induction and their effects on the chitinase biosynthesis were investigated. The RMS analysis suggested that the optimal conditions for production of recombinant ChiA are at lactose concentration of 6.15 mM, 8.12 hours after induction and cell density OD600nm = 0.87 with predicted enzyme activity of 7.49 U/ml. The model obtained was used for fermentation of recombinant strain and the real chitinase activity achieved 7.54 U/ml which is 1.32 times higher compared to that of the pre-optimization (5.71 U/ml). The RSM was found to be useful in optimizing and determining the interactions among process variables in ChiA enzyme production. Our data also indicated that using lactose as the inducer instead of, ITPG for ChiA expression can also reduce product costs.

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