Meyer-Barton E C, Klein J R, Imam M, Plapp R
Universität Kaiserslautern, Abteilung Mikrobiologie, Germany.
Appl Microbiol Biotechnol. 1993 Oct;40(1):82-9. doi: 10.1007/BF00170433.
Lactobacillus delbrückii ssp. lactis DSM7290 possesses an X-prolyl-dipeptidyl-aminopeptidase, designated PepX, which catalyses the hydrolytic removal of N-terminal dipeptidyl residues from peptides containing proline in the penultimate position. Using the specific substrate L-Ala-L-Pro-p-nitroanilide, PepX was purified by a four-step procedure including ammonium sulphate fractionation, hydrophobic interaction chromatography, ion exchange chromatography, and affinity chromatography. The N-terminus of the purified protein was sequenced. Screening of a gene library of chromosomal Lactobacillus delbrückii ssp. lactis DSM7290 DNA in the low-copy-number vector pLG339 resulted in the identification of the pepX gene in Escherichia coli using a specific plate assay with Gly-L-Pro-beta-naphthylamide as substrate. Nucleotide sequence analysis revealed an open reading frame of 2376 bp, coding for a protein of 792 amino acids with a molecular mass of 88449 Da.
德氏乳杆菌乳酸亚种DSM7290含有一种X-脯氨酰-二肽基氨基肽酶，命名为PepX，它能催化从倒数第二位含有脯氨酸的肽中水解去除N端二肽基残基。使用特异性底物L-丙氨酰-L-脯氨酰-对硝基苯胺，通过包括硫酸铵分级分离、疏水相互作用色谱、离子交换色谱和亲和色谱在内的四步程序对PepX进行纯化。对纯化蛋白的N端进行测序。以低拷贝数载体pLG339构建德氏乳杆菌乳酸亚种DSM7290染色体DNA的基因文库，通过以甘氨酰-L-脯氨酰-β-萘酰胺为底物的特异性平板检测，在大肠杆菌中鉴定出pepX基因。核苷酸序列分析揭示了一个2376 bp的开放阅读框，编码一个由792个氨基酸组成、分子量为88449 Da的蛋白质。
