Characterization of a novel potential peptide import system of Treponema denticola

Treponema denticola is a major etiologic agent of chronic periodontitis. On the outer sheath of T. denticola, several proteins such as the major outer sheath protein were detected, and among them, a 95 kDa protein has not yet been characterized. The aim of this study was to characterize the function of this 95 kDa protein. A gene encoding this 95 kDa protein (TDE_1072) of T. denticola was inactivated by homologous recombination. We compared growth curves between the TDE_1072 mutant and wild-type strains as well as differences in gene expression by DNA microarray analysis. Differentially expressed genes identified by microarray analysis were confirmed by quantitative reverse transcription-polymerase chain reaction. The proteins encoded by TDE_1072, TDE_1073, TDE_1074, TDE_1075, and TDE_1076 shared respective similarities to the substrate-binding domain (DppA) of an ABC-type dipeptide/oligopeptide/nickel transport system, and to the permease component (DppB and DppC) and ATPase component (DppD and DppF) of an ABC-type dipeptide/oligopeptide/nickel transport system.　Inactivation of dppA attenuated the growth of T. denticola and down-regulated expression of dppB-dppF. In contrast, expression of oppB-oppF was upregulated in the mutant. Our findings indicate that TDE_1072 may be a potential periplasmic solute binding protein encoded by dppA that is involved in the organization of a peptide uptake system with dppB-dppF. Comparison of 2 strains of T. denticola. Treponema denticola ATCC 35405 and I-1 was cultured in TYGVS medium for 24-48 h. Profile of gene expression of T. denticola I-1 was compared to that of ATCC 35405.

齿垢密螺旋体（Treponema denticola）是慢性牙周炎的主要致病因子。在该菌的外鞘上已检测到包括主要外鞘蛋白在内的多种蛋白，其中一种分子量为95 kDa的蛋白尚未得到功能表征。
本研究旨在解析该95 kDa蛋白的功能。研究人员通过同源重组（homologous recombination）技术，失活了齿垢密螺旋体中编码该95 kDa蛋白的基因（TDE_1072）。
随后，对比了TDE_1072突变株与野生型菌株的生长曲线，并通过DNA微阵列分析（DNA microarray analysis）检测两组的基因表达差异。通过微阵列分析筛选得到的差异表达基因，经定量逆转录聚合酶链式反应（quantitative reverse transcription-polymerase chain reaction）进行了验证。
TDE_1072、TDE_1073、TDE_1074、TDE_1075及TDE_1076所编码的蛋白，分别与ABC型二肽/寡肽/镍转运系统（ABC-type dipeptide/oligopeptide/nickel transport system）的底物结合结构域（DppA）、通透酶组分（DppB、DppC）以及ATP酶组分（DppD、DppF）具有序列相似性。
dppA基因的失活会减弱齿垢密螺旋体的生长，并下调dppB-dppF的基因表达；与之相反，突变株中oppB-oppF的基因表达则出现上调。
本研究结果表明，TDE_1072可能是dppA编码的潜在周质溶质结合蛋白，参与由dppB-dppF构成的肽摄取系统的组装。
对两株齿垢密螺旋体进行比较实验：将ATCC 35405株与I-1株齿垢密螺旋体在TYGVS培养基中培养24~48小时，对比I-1株与ATCC 35405株的基因表达谱。