酰胺化酶基因及其表达活性产物对多肽的酰胺化修饰应用

摘要

本发明涉及一种大鼠来源的酰胺化酶基因及其在仓鼠卵巢细胞（ＣＨＯ）的活性表达产物。通过原位杂交及ＰＣＲ方法，从大鼠脑ｃＤＮＡ库中，筛选到３个大鼠甘氨肽酰化单氧酶（ｒＰＡＭ）基因片段，经点突变、ＰＣＲ重组技术，拼接出ｒＰＡＭ全酶基因，并构建其真核表达质粒，在ＣＨＯ细胞中获得活性表达，表达产物可直接用于多肽或蛋白的酰胺化修饰。

主权项

1、一种重组大鼠酰胺化酶基因，其特征在于该重组基因具有以下DNA核苷酸序列：CTGCGAATTC 10ATGGCCGGACGCGCCCGCAGCGGTCTGCTACTGCTGCTGCTGGGGCTGCTCGCCCTGCAG 70AGCAGCTGCCTGGCCTTCAGAAGCCCACTTTCTGTCTTTAAGAGGTTTAAAGAAACTACC 130AGATCATTTTCCAATGAATGCCTTGGTACCATTGGACCAGTCACCCCTCTTGATGCATCA 190GATTTTGCGCTGGATATTCGCATGCCTGGGGTTACACCTAAAGAGTCTGACACATACTTC 250TGCATGTCCATGCGTCTGCCTGTGGATGAGGAAGCCTTCGTGATTGACTTCAAGCCTCGT 310GCCAGCATGGATACTGTCCACCATATGCTGCTGTTTGGATGCAATATGCCCTCGTCCACT 370GGAAGTTACTGGTTTTGTGATGAAGGAACCTGTACAGATAAAGCCAATATTCTATATGCC 430TGGGCAAGGAATGCTCCCCCCACCCGGCTCCCGAAAGGTGTTGGATTCAGAGTTGGAGGA 490GAAACTGGAAGCAAATACTTCGTCCTTCAAGTTCACTATGGCGATATCAGTGCTTTTCGA 550GATAATCACAAAGACTGCTCTGGCGTGTCCGTACATCTCACACGTGTGCCCCAGCCTTTA 610ATTGCGGGCATGTACCTTATGATGTCTGTTGACACTGTCATACCACCAGGAGAGAAAGTA 670GTGAATGCTGACATTTCGTGCCAATACAAAATGTATCCAATGCATGTGTTTGCCTACAGA 730GTCCACACTCACCATTTAGGTAAGGTGGTGAGCGGATACAGAGTAAGAAACGGACAGTGG 790ACACTGATTGGACGCCAGAACCCCCAGCTGCCACAGGCTTTCTACCCTGTGGAACACCCC 850GTTGATGTTACTTTTGGTGATATACTGGCAGCCAGATGTGTGTTCACTGGTGAAGGGAGG 910ACAGAGGCCACCCATATCGGCGGCACTTCTAGTGACGAAATGTGTAACCTGTACATCATG 970TATTACATGGAAGCCAAATATGCACTTTCCTTCATGACCTGTACAAAGAACGTGGCTCCA 1030GATATGTTCAGAACTATCCCAGCAGAGGCCAATATCCCAATTCCTGTCAAACCGGACATG 1090GTTATGATGCACGGGCATCACAAAGAAGCAGAAAACAAAGAAAAGAGTGCTTTAATGCAG 1150CAGCCAAAACAGGGAGAGGAAGAAGTATTAGAGCAGGATCCTTTCCATGTGGAAGAAGAA 1210CTGGACTGGCCTGGAGTGTACTTGTTACCAGGCCAGGTTTCTGGGGTGGCCCTGGATTCT 1270AAGAATAACCTAGTGATTTTCCACAGAGGTGACCATGTTTGGGATGGAAACTCTTTTGAC 1330AGCAAGTTTGTTTACCAGCAAAGAGGTCTTGGGCCAATTGAAGAAGACACCATCCTGGTC 1390ATTGACCCAAATAATGCTGAAATCCTCCAGTCCAGTGGCAAGAACCTGTTTTATTTACCA 1450CACGGCTTGAGCATAGATACAGATGGAAATTATTGGGTCACAGATGTGGCTCTCCACCAG 1510GTGTTCAAATTGGACCCGCATAGCAAAGAAGGCCCTCTCTTAATTCTGGGAAGGAGCATG 1570CAACCTGGGAGTGACCAAAATCATTTCTGCCAGCCCACCGATGTGGCTGTGGAGCCCAGT 1630ACTGGAGCTGTCTTCGTGTCAGACGGTTACTGTAACAGTCGGATTGTGCAGTTTTCACCA 1690AGCGGAAAGTTCGTCACCCAGTGGGGAGAAGAGTCCTCTGGAAGCAGTCCTAGGCCAGGC 1750CAGTTCAGTGTTCCTCACAGTTTGGCCCTTGTGCCTCATTTGGACCAGTTGTGTGTGGCA 1810GACAGGGAAAATGGCCGAATCCAATGCTTCAAAACTGACACCAAAGAATTTGTGAGAGAG 1870ATTAAGCACGCATCATTTGGAAGGAATGTCTTTGCCATTTCATATATACCAGGTTTCCTC 1930TTTGCCGTAAACGGGAAGCCTTACTTTGGAGACCAAGAGCCCGTGCAAGGATTTGTGATG 1990AACTTTTCCAGTGGGGAAATTATAGACGTCTTCAAGCCAGTACGCAAGCACTTCGACATG 2050CCTCATGATATTGTGGCTTCTGAAGATGGGACTGTGTACATTGGAGACGCACACACAAAC 2110ACCGTGTGGAAGTTCACCCTGACTGAAAAAATGGAGCATCGGTCAGTTAAAAAGGCTGGC 2170ATTGAAGTCCAGGAAATCAAAGAAGCCGAGGCAGTTGTTGAACCCAAAGTGGAGAACAAA 2230CCCACCTCCTCAGAATTGCAGAAGATGCAAGAGAAACAGAAACTGAGCACAGAGCCCGGC 2290TCGGGAGTGTCCGTGGTTCTCATTACAACCCTTCTGGTTATTCCTGTGCTGGTCCTGCTG 2350GCCATTGTCATGTTTATTCGGTGGAAAAAATCAAGGGCCTTTGGAGATCATGACCGCAAG 2410CTCGAGTCAAGTTCTGGAAGAGTCCTGGGAAGATTCCGAGGAAAGGGAAGCGGCGGCTTA 2470AATCTGGGAAATTTCTTTGCAAGTCGAAAAGGCTACAGCAGAAAAGGGTTTGACCGAGTG 2530AGCACAGAGGGGAGTGACCAAGAGAAAGATGAGGACGACGGAACTGAGTCTGAAGAGGAG 2590TACTCGGCCCCGCTGCCCAAGCCTGCACCTTCCTCCTGA 2629GCCTCCAGCCTTCGCCCGGGTAGCTGGACTGAGGTTTACCAGGATGCCCAGACTCCTTCC 2689CCTTTAGCGCGTGTAAAGTTCTGTGCATTTGATTGTAAACTGTACTCGTCAGTGTGGGAC 2749TGTACACACCTTATTTACTTCATTTGGCTCCGTTGGCTTCTGTTCTCTAGGTGAGGAGTT 2809CCCCACCAGTTCACTCCAGTGCCATTGTCTTTATATGAACTTAGCGTAGAGAAGCCGCCC 2869TCCTCTTCCAAGGTAGCGCTCCAACCCCCGAGGGAAGTTTAGCTCATTCACATTTGGAGA 2929CGTTTTAGTTGGTGGATGTAAATAGCCCTATTCTCTGCTTGAACACAGTATTCTCCCAGT 2989CCACACCCATCGCCAGTGTCTTTCTTTGGTGCCTTTCCTGTTCAGCATTCTCAGCCTGTG 3049GCAGTGAAGAGAACCAACCTGCCACACGACGAAAAGCTGCTAAATCTCCTTCTATTTTTT 3109TAAAATCACTAACATTATATTGCAATGAGAGAAATTTTAAAAAGTCTCTATTTAAATTCT 3169TTTTTTAAATTTCTCCTCAGTTGGTGTGTTTCCGCTCGTGCCGAATTC 3217