Takahashi M, Pischetsrieder M, Monnier V M
Institute of Pathology, School of Medicine, Case Western Reserve University, Cleveland, Ohio 44106, USA.
J Biol Chem. 1997 May 9;272(19):12505-7. doi: 10.1074/jbc.272.19.12505.
Amadoriase is an enzyme catalyzing the oxidative deglycation of Amadori products to yield corresponding amino acids, glucosone, and H2O2. We previously reported the purification and characterization of two amadoriase isozymes from Aspergillus sp. that degrade both glycated low molecular weight amines and amino acids (Takahashi, M., Pischetsrieder, M., and Monnier, V. M. (1997) J. Biol. Chem. 272, 3437-3443). To identify the primary structure of the enzymes, we have prepared a cDNA library from Aspergillus fumigatus induced with fructosyl propylamine and isolated a clone using polyclonal anti-amadoriase II antibody. The primary structure of the enzyme deduced from the nucleotide sequence comprises 438 amino acid residues with a predicted molecular mass of 48,798 Da. The deduced primary structure exhibits the presence of an ADP-binding motif near the NH2 terminus. The identity of the amadoriase II cDNA was further confirmed by expression in Escherichia coli cells with an inducible expression system. Northern blotting analysis revealed that amadoriase II was induced by fructosyl propylamine in a dose-dependent manner.
氨基脱氧酮糖酶是一种催化氨基脱氧酮糖产物氧化脱糖基化反应生成相应氨基酸、葡萄糖酮和过氧化氢的酶。我们之前报道过从曲霉属真菌中纯化和鉴定出两种氨基脱氧酮糖酶同工酶，它们能降解糖化的低分子量胺和氨基酸（高桥，M.，皮谢茨里德，M.，和莫尼尔，V. M.（1997年）《生物化学杂志》272，3437 - 3443）。为了确定这些酶的一级结构，我们用果糖基丙胺诱导烟曲霉制备了一个cDNA文库，并使用多克隆抗氨基脱氧酮糖酶II抗体分离出一个克隆。从核苷酸序列推导的酶的一级结构包含438个氨基酸残基，预测分子量为48,798道尔顿。推导的一级结构在NH2末端附近显示存在一个ADP结合基序。通过在大肠杆菌细胞中用诱导表达系统进行表达，进一步证实了氨基脱氧酮糖酶II cDNA的身份。Northern印迹分析表明，果糖基丙胺以剂量依赖的方式诱导氨基脱氧酮糖酶II。
