介孔二氧化钛固载γ-谷氨酰转肽酶的制备及性质  

Immobilization of GGT on meso-TiO_2:preparation and properties

作　　者：秦丹华[1] 姚忠[1] 王浩琦[1] 肖易凡[1] 杨敬[1] 徐虹[1] 韦萍[1] 

机构地区：[1]南京工业大学食品与轻工学院,江苏南京210009

出　　处：《化工学报》2011年第2期378-385,共8页CIESC Journal

基　　金：国家重点基础研究发展计划项目(2009CB724706);江苏省高校自然科学研究重大项目(10KJA530014)~~

摘　　要：γ-谷氨酰转肽酶(GGT)在临床诊断和生物催化方面具有重要的应用价值。本文以介孔氧化钛晶须为载体进行GGT的固定化,考察了载体结构特性、吸附时间和给酶量对固定化效果的影响,并对固定化酶的催化特性及其稳定性进行了研究。结果显示,以最可几孔径为30nm的介孔TiO2为载体,载体载酶量可达5.07mg·g-1。在给酶量为18.99 U·g-1时,经室温吸附2.5h,固定化酶活性回收率可达73.05%。固定化酶的pH稳定性和热稳定性均显著优于游离酶,在4℃下保温贮藏60d、转化22个批次后,固定化酶活力仍可保持初始值的71.30%。经测定,游离酶和固定化酶的米氏常数Km分别为0.79mmol·L-1和1.05mmol.L-1,酰基化反应活化能分别为13.59 kJ·mol-1和15.42 kJ·mol-1;固定化GGT的失活反应活化能Ed为92.80 kJ·mol-1,相比于游离酶(49.61 kJ·mol-1)有明显的增加。γ-Glutamyltranspeptidase（GGT）is an important enzyme with wide applications in biocatalysis and clinical diagnosis.In this work,mesoporous fibrous titania（M-TiO2）was used for immobilization of GGT from B.subtilis NX-2 and the properties of immobilized GGT were also investigated.When the M-TiO2 support with average pore diameter of 30 nm was used,the amount of immobilized protein was 5.07 mg·g-1,and the yield of activity was 73.05% at the ratio of GGT/support was 18.99 U·g-1 after incubation at room temperature for 2.5 h.The thermal and pH stability of the immobilized GGT was higher than that in its free form.After storage at 4℃ for 60 days and repeated use for 22 batches,the activity of the immobilized GGT remained 71.30% of its initial activity.The kinetic parameters（Km）for free and immobilized GGT were determined as 0.79 mmol·L-1 and 1.05 mmol·L-1,respectively.The activation energy（Ea）values of glutamylation were 15.42 kJ·mol-1and 13.59 kJ·mol-1 for immobilized and free GGT.The thermal inactivation energy（Ed）values of GGT for immobilized and free enzyme were also calculated to be 92.80 kJ·mol-1 and 49.61 kJ·mol-1,respectively.

关 键 词：Γ-谷氨酰转肽酶 固定化 介孔二氧化钛 酶学性质 

分 类 号：TQ936.2[化学工程]