Hagting A, Kunji E R, Leenhouts K J, Poolman B, Konings W N
Department of Microbiology, University of Groiningen, Haren, The Netherlands.
J Biol Chem. 1994 Apr 15;269(15):11391-9.
Lactococcus lactis takes up di- and tripeptides via a proton motive force-dependent carrier protein. The gene (dtpT) encoding the di-tripeptide transport protein of L. lactis was cloned by complementation of a dipeptide transport-deficient and proline auxotrophic Escherichia coli strain. Functional expression of the dipeptide transport gene was demonstrated by uptake studies of alanyl-[14C]glutamate and other peptides in E. coli cells. The di-tripeptide transport protein catalyzes proton motive force-driven peptide uptake and dipeptide exchange activity. The nucleotide sequence of dtpT was determined and the translated sequence corresponds with a protein of 463 amino acid residues. Hydropathy profiling indicates that the protein could form 12 membrane-spanning segments with the amino and carboxyl termini at the outer surface of the membrane. A secondary structure model is presented which is substantiated by analysis of DtpT-PhoA fusion constructs. Amino acid sequence comparisons showed no significant homology with other bacterial peptide transport systems nor with any other known protein. Flanking regions of the di-tripeptide transport gene were used to delete dtpT from the chromosome of L. lactis. Genetic and biochemical characterization of this mutant indicates that DtpT is the only transport protein in L. lactis for hydrophilic di- and tripeptides.
乳酸乳球菌通过一种质子动力依赖型载体蛋白摄取二肽和三肽。通过对一株缺乏二肽转运能力且脯氨酸营养缺陷型的大肠杆菌菌株进行互补，克隆出了编码乳酸乳球菌二肽转运蛋白的基因（dtpT）。通过对大肠杆菌细胞中丙氨酰-[14C]谷氨酸和其他肽的摄取研究，证实了二肽转运基因的功能表达。二肽 - 三肽转运蛋白催化质子动力驱动的肽摄取和二肽交换活性。测定了dtpT的核苷酸序列，其翻译后的序列对应于一个由463个氨基酸残基组成的蛋白质。亲水性分析表明，该蛋白可形成12个跨膜区段，其氨基和羧基末端位于膜的外表面。本文提出了一个二级结构模型，该模型通过对DtpT - PhoA融合构建体的分析得到了证实。氨基酸序列比较显示，该蛋白与其他细菌肽转运系统以及任何其他已知蛋白均无明显同源性。利用二肽 - 三肽转运基因的侧翼区域从乳酸乳球菌染色体中删除了dtpT。对该突变体的遗传和生化特性分析表明，DtpT是乳酸乳球菌中唯一负责亲水性二肽和三肽转运的蛋白。
