Bretting H, Jacobs G
Biochim Biophys Acta. 1987 Jul 7;913(3):342-8. doi: 10.1016/0167-4838(87)90145-2.
The enzymic oxidation of snail galactans, of their first and second Smith degradation products and of some structurally related polysaccharides was studied. Lymnaea stagnalis galactan after one cycle of Smith degradation reacted best and native Helix pomatia galactan was almost inactive. Investigations on the structural requirements for oligosaccharides to bind to galactose oxidase showed that the branched tetrasaccharide, Gal-beta-1----6-[Gal-beta-1----3]-Gal-beta-1----1 L-Gro, in the terminal nonreducing position was the most complementary structure in the native galactan to associate with the enzyme. All nitrophenyl alpha-galactosides reacted better, and the ortho-form was 10-times more potent compared with this tetrasaccharide, indicative of the involvement of a hydrophobic region in binding. However, the beta-linked isomers were only equally or less reactive than galactose. The enzymic oxidation determined colorimetrically by transferring the peroxide formed to o-dianisidine ceased at a maximum typical for each substrate and independent of the reaction time. When the absolute turn-over rates for ortho- and para-nitrophenyl alpha-galactoside and for the beta-isomer were determined by HPLC, it could be demonstrated that the oxidation had not finished at the maximum of the colour reaction, but proceeded until the substrate was consumed. The initial speed of the colour reaction paralleled the absolute oxidation rate.
研究了蜗牛半乳聚糖、其一级和二级史密斯降解产物以及一些结构相关多糖的酶促氧化反应。椎实螺半乳聚糖经过一轮史密斯降解后反应最佳，而天然的苹果螺半乳聚糖几乎没有活性。对寡糖与半乳糖氧化酶结合的结构要求的研究表明，在末端非还原位置的分支四糖Gal-β-1----6-[Gal-β-1----3]-Gal-β-1----1 L-Gro是天然半乳聚糖中与该酶结合最互补的结构。所有对硝基苯基α-半乳糖苷反应更好，邻位形式的活性比该四糖高10倍，这表明结合过程中涉及一个疏水区域。然而，β-连接的异构体的反应活性仅与半乳糖相当或更低。通过将生成的过氧化物转移至邻联茴香胺进行比色测定的酶促氧化反应在每种底物的典型最大值处停止，且与反应时间无关。当通过高效液相色谱法测定邻硝基苯基α-半乳糖苷、对硝基苯基α-半乳糖苷和β-异构体的绝对周转速率时，可以证明氧化反应在显色反应最大值时并未完成，而是持续进行直至底物消耗殆尽。显色反应的初始速度与绝对氧化速率平行。
