细菌脱色酶TpmD对三苯基甲烷类染料脱色的酶学特性研究  

Properties of a triphenylmethane dyes decolorization enzyme TpmD from Aeromonas hydrophila strain DN322

作　　者：任随周[1] 郭俊[1] 王亚丽[1] 岑英华[1] 孙国萍[1] 

机构地区：[1]广东省微生物研究所广东省菌种保藏与应用重点实验室

出　　处：《微生物学报》2006年第3期385-389,共5页Acta Microbiologica Sinica

基　　金：国家863项目(2003AA214040);广东省自然科学基金团队项目(015017);广东省科技攻关重大项目(2004A30404002);广东省自然科学基金项目(032319)~~

摘　　要：从嗜水气单胞菌DN322中分离纯化出能够对三苯基甲烷类染料结晶紫、碱性品红、灿烂绿及孔雀绿进行有效脱色的脱色酶，命名为TpmD。该酶的亚基分子量为29．4kDa，等电点为5．6。该酶催化上述4种三苯基甲烷类染料脱色反应的适合温度为40～60℃，适合pH范围为5．5～9．0。动力学参数测定结果显示TpmD对结晶紫、碱性品红、灿烂绿及孔雀绿的Km值分别为24．3、40．6、54．2、68．5μmol^-1·L^-1，Vmax值分别为19．6、74．1、82．8、115．6μmol^-1·L^-1。结晶紫为该酶的最适反应底物。TpmD催化的脱色反应依懒于NADH／NADPH及分子氧的存在，显示该酶属于NADH／NADPH依赖型的氧化酶类。这是国内外首次关于细菌中三苯基甲烷类染料脱色酶酶学性质的描述。A novel bacterial decolorization enzyme for triphenylmethane dyes from Aeromonas hydrophila strain DN322 was purified, named TpmD. The purified enzyme catalyzes the decolorization of several triphenylmethane dyes, i.e., crystal violet, basic fuchsin, brilliant green and malachite green. The enzyme was identified by the clear transparent band development of zymogram stained with crystal violet, basic fuchsin, brilliant green and malachite green after polyacrylamide gel electrophoresis （PAGE） respectively. The decolorization enzyme was enzymologically characterized. The results showed that the molecular weight of TpmD is 29.4kDa and its isoelectric point （pt） is 5.6. The maximal activity of TpmD for above four triphenylmethane dyes was observed at 50℃- 55℃ and pH 7.4 ～ 8.0. The temperature for losing half of the activity （t1/2） within 4h is 62℃. The activities of decolorization enzyme are relatively stable at pH range of 5.5 - 9.0. The Km and Vmax of TpmD for decolorizing crystal violet, basic fuchsin, brilliant green and malachite green are 24.3,40.6,54.2,68.5μmol/L respectively, Vmax are 19.6,74.1,82.8,115,6μmol^-1·L^-1·s^-1 respectively. Both NADH/NADPH and molecular oxygen are necessary for the enzyme to decolorize triphenylmethane dyes, indicate the enzyme is an NADH/NADPHdependent oxygenase.

关 键 词：脱色酶 嗜水气单胞菌 三苯基甲烷类染料 酶学特性 

分 类 号：X172[环境科学与工程—环境科学]