Beijing Key Laboratory of Bioprocess, College of Life Science and Technology, Beijing University of Chemical Technology, Beijing 100029, People's Republic of China.
Center for Synthetic and Systems Biology, School of Life Sciences, Tsinghua University, Beijing 100084, China.
Int J Biol Macromol. 2024 Mar;261(Pt 2):129838. doi: 10.1016/j.ijbiomac.2024.129838. Epub 2024 Feb 1.
A novel α-amylase Amy03713 was screened and cloned from the starch utilization strain Vibrio alginolyticus LHF01. When heterologously expressed in Escherichia coli, Amy03713 exhibited the highest enzyme activity at 45 °C and pH 7, maintained >50 % of the enzyme activity in the range of 25-75 °C and pH 5-9, and sustained >80 % of the enzyme activity in 25 % (w/v) of NaCl solution, thus showing a wide range of adapted temperatures, pH, and salt concentrations. Halomonas bluephagenesis harboring amy03713 gene was able to directly utilize starch. With optimized amylase expression, H. bluephagenesis could produce poly(3-hydroxybutyrate) (PHB), poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV), and poly(3-hydroxybutyrate-co-4-hydroxybutyrate) (P34HB). When cultured for PHB production, recombinant H. bluephagenesis was able to grow up to a cell dry weight of 11.26 g/L, achieving a PHB titer of 6.32 g/L, which is the highest titer that has been reported for PHB production from starch in shake flasks. This study suggests that Amy03713 is an ideal amylase for PHA production using starch as the carbon source in H. bluephagenesis.
从淀粉利用菌弧菌 LHF01 中筛选并克隆了一种新型的α-淀粉酶 Amy03713。当在大肠杆菌中异源表达时，Amy03713 在 45°C 和 pH 7 下表现出最高的酶活性，在 25-75°C 和 pH 5-9 的范围内保持>50%的酶活性，在 25%（w/v）的 NaCl 溶液中维持>80%的酶活性，因此显示出广泛的适应温度、pH 值和盐浓度范围。携带 amy03713 基因的嗜盐菌能够直接利用淀粉。通过优化淀粉酶的表达，H. bluephagenesis 可以生产聚（3-羟基丁酸酯）（PHB）、聚（3-羟基丁酸酯-3-羟基戊酸酯）（PHBV）和聚（3-羟基丁酸酯-4-羟基丁酸酯）（P34HB）。当用于生产 PHB 时，重组 H. bluephagenesis 能够生长到 11.26 g/L 的细胞干重，实现 6.32 g/L 的 PHB 产量，这是在摇瓶中报道的利用淀粉生产 PHB 的最高产量。本研究表明，Amy03713 是利用淀粉作为碳源在嗜盐菌中生产 PHA 的理想淀粉酶。
