State Key Laboratory of Food Science and Technology, School of Food Science and Technology, Jiangnan University, 1800 Lihu Road, Wuxi 214122, China; Department of Food Science and Technology, Faculty of Engineering and Technology, University of Gezira, P.O. Box 20, Wad Madani, Sudan.
Ultrason Sonochem. 2014 Jan;21(1):76-83. doi: 10.1016/j.ultsonch.2013.04.014. Epub 2013 May 9.
In our current research work, the effect of combination of ultrasonic irradiation and high hydrostatic pressure (US/HHP) on the enzymatic activity and enzymatic hydrolysis kinetic parameters of dextran catalytic by dextranase were investigated. Furthermore, the effects of US/HHP on the structure of dextranase were also discussed with the aid of fluorescence spectroscopy and circular dichroism (CD) spectroscopy. The maximum hydrolysis of dextran was observed under US (40 W at 25 kHz for 15 min) combined with HHP (400 MPa for 25 min), in which the hydrolysis of dextran increased by 163.79% compared with the routine thermal incubation at 50 °C. Results also showed that, Vmax and KM values, as well as, kcat of dextranase under US/HHP treatment were higher than that under US, HHP and thermal incubation at 50 °C, indicated that, the substrate is converted into the product at an increased rate when compared with the incubation at 50 °C. Compared to the enzymatic reaction under US, HHP, and routine thermal incubation, dextranase enzymatic reaction under US/HHP treatment showed decreases in Ea, ΔG and ΔH, however small increase in ΔS value was observed. In addition, fluorescence and CD spectra reflected that US/HHP treatment had increased the number of tryptophan on dextranase surface with increased α-helix by 19.80% and reduced random coil by 6.94% upon US/HHP-treated dextranase protein compared to the control, which were helpful for the improvement of its activity. These results indicated that, the combination of US and HHP treatments could be an effective method for improving the hydrolysis of dextran in many industrial applications including sugar manufacturing processes.
在我们目前的研究工作中，研究了超声辐射和高静压（US/HHP）联合作用对葡聚糖酶催化的葡聚糖酶酶活性和酶水解动力学参数的影响。此外，还借助荧光光谱和圆二色性（CD）光谱讨论了 US/HHP 对葡聚糖酶结构的影响。在 US（25 kHz 下 40 W 超声 15 分钟）与 HHP（400 MPa 高压 25 分钟）联合作用下，观察到葡聚糖的最大水解，与常规 50°C 热孵育相比，葡聚糖的水解增加了 163.79%。结果还表明，在 US/HHP 处理下，Vmax 和 KM 值以及 kcat 均高于 US、HHP 和 50°C 热孵育，表明与 50°C 孵育相比，底物以更高的速率转化为产物。与 US、HHP 和常规热孵育下的酶反应相比，US/HHP 处理下的葡聚糖酶反应表现出 Ea、ΔG 和 ΔH 的降低，而 ΔS 值略有增加。此外，荧光和 CD 光谱反映出，与 US、HHP 和常规热孵育下的酶反应相比，US/HHP 处理后，葡聚糖酶表面的色氨酸数量增加，α-螺旋增加了 19.80%，无规卷曲减少了 6.94%，这有助于提高其活性。这些结果表明，US 和 HHP 联合处理可以成为提高许多工业应用中（包括糖制造过程）葡聚糖水解的有效方法。
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