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以辣根过氧化物酶(horseradish peroxidase,HRP)为目标蛋白,N-异丙基丙烯酰胺(NIPAM)为单体,采用原位沉淀聚合法,在37℃,NIPAM和HRP质量比为6∶1的条件下,制备了粒径大小为13.7 nm,zeta电位是(-3.7±0.3)mV的温度响应型辣根过氧化物酶纳米胶囊(nHRP).采用基质辅助激光解吸电离飞行时间质谱仪(MALDI-TOF MS)、透射电镜(TEM)、动态光散射仪(DLS)和紫外-可见光分光光度计(UV-Vis)等对HRP的修饰度,nHRP的微观结构、粒径大小、催化活性、温度响应性能和热稳定性能进行了表征.研究表明,制备的辣根过氧化物酶纳米胶囊单分散性较好;在环境温度高于33℃时,nHRP出现响应变化,其粒径大小变化显著,且呈可逆性;nHRP相比HRP原酶热稳定性显著提高;50℃时,将nHRP高速离心,可使酶多次有效地分离和回收.
Using horseradish peroxidase (HRP) as target protein and N-isopropylacrylamide (NIPAM) as monomer, the in situ precipitation polymerization was used at 37 ℃, the mass ratio of NIPAM to HRP was 6: (NHRP) with a size of 13.7 nm and a zeta potential of (-3.7 ± 0.3) mV, was prepared by using a matrix-assisted laser desorption / ionization time-of-flight mass spectrometer (MALDI-TOF MS), transmission electron microscopy (TEM), dynamic light scattering (DLS) and ultraviolet-visible spectrophotometer (UV-Vis) on the degree of modification of HRP, the microstructure of nHRP, particle size, catalytic activity , Temperature response performance and thermal stability were characterized.The results showed that the preparation of horseradish peroxidase nanocapsules monodisperse better; at ambient temperatures above 33 ℃, nHRP response changes, the size of the particle size changes Significantly, and was reversible; nHRP compared HRP protenzyme thermal stability was significantly improved; 50 ℃, the nHRP high-speed centrifugation, the enzyme can be effectively separated and recovered many times.