Ugarova N N, Rozhkova G D, Berezin I V
Biokhimiia. 1978 Jul;43(7):1242-50.
Effect of chemical modification of horseradish peroxidase lysine epsilon-amino groups by propionic, butyric, valeric, succinic anhydrides and trinitrobenzolsulfonic acid (TNBS) on catalytic properties of the enzyme is investigated. All the preparations of modified peroxidase have 100% peroxidase activity for o-dianizidine at pH 7.0, which indicates the absence of lysine epsilon-amino group in the enzyme active site. pH-dependencies of modified peroxidase relative activity are studied; modification by anhydrides of monobasic acids is not found to result in changes of the relative activity pH-profile, while modification by succinic anhydride widens it. Absorption and circular dichoism spectra of native and modified peroxidase within 260--270 nm are obtained, some changes in the enzyme tertiary structure after its epsilon-amino groups modification are observed. Modification of four epsilon-amino groups by buturic and succinic anhydrides and of three epsilon-amino groups by TNBS is found to increase the regidity of protein surrounding of heme, and modification of six epsilon-amino groups by TNBS results in more unwrapped enzyme structure as compared with its native molecule.
研究了用丙酸酐、丁酸酐、戊酸酐、琥珀酸酐和三硝基苯磺酸（TNBS）对辣根过氧化物酶赖氨酸ε-氨基进行化学修饰对该酶催化特性的影响。所有修饰过的过氧化物酶制剂在pH 7.0时对邻联茴香胺均具有100%的过氧化物酶活性，这表明酶活性位点中不存在赖氨酸ε-氨基。研究了修饰过的过氧化物酶相对活性的pH依赖性；发现一元酸酐修饰不会导致相对活性pH曲线发生变化，而琥珀酸酐修饰会使其变宽。获得了天然和修饰过的过氧化物酶在260 - 270 nm范围内的吸收光谱和圆二色光谱，观察到酶ε-氨基修饰后其三级结构有一些变化。发现用丁酸酐和琥珀酸酐修饰四个ε-氨基以及用TNBS修饰三个ε-氨基会增加血红素周围蛋白质的刚性，与天然分子相比，用TNBS修饰六个ε-氨基会导致酶结构更加松散。
