Hirokawa Kozo, Gomi Keiko, Kajiyama Naoki
Research and Development Division, Kikkoman Corporation, Noda-city, Chiba Pref. 278-0037, Japan.
Biochem Biophys Res Commun. 2003 Nov 7;311(1):104-11. doi: 10.1016/j.bbrc.2003.09.169.
Fructosyl peptide oxidases, enzymes that are active against a model compound of glycated hemoglobin, N(alpha)-fructosyl valyl-histidine, were characterized. To identify the primary structure of fructosyl peptide oxidases, we have prepared cDNA libraries from Eupenicillium terrenum ATCC18547 and Coniochaeta sp. NISL9330. The coding regions, both fungal fructosyl peptide oxidases consisting of 1314-bp, were obtained with degenerated primers based on the amino acid sequences and specific primers by 3(') and 5(') RACE (rapid amplification of cDNA ends). By their sequence similarities and substrate specificities, fructosyl peptide oxidases and their homologs could be categorized into two groups: (A) enzymes that preferably oxidize alpha-glycated molecules and (B) enzymes that preferably oxidize epsilon-glycated molecules. We showed that recombinant fructosyl peptide oxidases could be used to detect protease-treated fructosyl-hexapeptide, a glycated peptide that is released from HbA(1C) by endoproteinase Glu-C, suggesting these enzymes could be useful for the enzymatic measurement of HbA(1C).
对果糖基肽氧化酶进行了表征，该酶对糖化血红蛋白的模型化合物N(α)-果糖基缬氨酰-组氨酸具有活性。为了确定果糖基肽氧化酶的一级结构，我们从地青霉ATCC18547和锥毛壳菌属NISL9330制备了cDNA文库。基于氨基酸序列使用简并引物，并通过3(')和5(')RACE（cDNA末端快速扩增）使用特异性引物，获得了两种真菌果糖基肽氧化酶均由1314 bp组成的编码区。根据它们的序列相似性和底物特异性，果糖基肽氧化酶及其同源物可分为两组：(A) 优先氧化α-糖化分子的酶和 (B) 优先氧化ε-糖化分子的酶。我们表明重组果糖基肽氧化酶可用于检测蛋白酶处理的果糖基六肽，这是一种由内肽酶Glu-C从HbA(1C)释放的糖化肽，表明这些酶可用于HbA(1C)的酶促测定。
