Villette J R, Helbecque N, Albani J R, Sicard P J, Bouquelet S J
Laboratoire de Chimie Biologique, Université des Sciences et Technologies de Lille, Villeneuve d'Ascq, France.
Biotechnol Appl Biochem. 1993 Apr;17(2):205-16.
Nitration of tyrosine residues was performed on Bacillus circulans E 192 cyclomaltodextrin glucanotransferase (CGTase) using tetranitromethane (TNM). A maximum of 15 out of 28 tyrosine residues is modified with 8 mM TNM, entailing a concomitant loss of enzymic activity and tryptophan fluorescence. Spectroscopic studies suggest that these two phenomena are related to an impairment of the enzyme conformation as a consequence of the tyrosine nitration. The presence of 5 mM acarbose during the CGTase nitration results in the protection of one tyrosine residue and the rate of inactivation is reduced 9.4-fold. These results support a contribution of a tyrosine residue in the CGTase catalytic site. The nitration of CGTase also entails a decrease in the enzyme's affinity for a beta-cyclodextrin (beta-CD) co-polymer. Kinetic and analytical investigations on isolated modified enzymes support the concept that this phenomenon is unrelated to the modification of tyrosine residues, but rather concerns a side reaction of the reagent occurring at the raw-starch-binding site of the CGTase.
使用四硝基甲烷（TNM）对环状芽孢杆菌E 192环麦芽糊精葡糖基转移酶（CGTase）的酪氨酸残基进行硝化。在28个酪氨酸残基中，最多有15个被8 mM的TNM修饰，这伴随着酶活性和色氨酸荧光的同时丧失。光谱研究表明，这两种现象与酪氨酸硝化导致的酶构象受损有关。在CGTase硝化过程中存在5 mM的阿卡波糖，可保护一个酪氨酸残基，使失活速率降低9.4倍。这些结果支持了一个酪氨酸残基在CGTase催化位点中的作用。CGTase的硝化还导致该酶对β-环糊精（β-CD）共聚物的亲和力降低。对分离出的修饰酶进行的动力学和分析研究支持了这样一种观点，即这种现象与酪氨酸残基的修饰无关，而是涉及试剂在CGTase的生淀粉结合位点发生的副反应。
