Burne R A, Parsons D T, Marquis R E
Department of Microbiology, University of Rochester School of Medicine and Dentistry, New York 14642.
Infect Immun. 1989 Nov;57(11):3540-8. doi: 10.1128/iai.57.11.3540-3548.1989.
The common oral bacterium Streptococcus sanguis can degrade arginine via the arginine deiminase (AD) system. The three enzymes of this system, AD, ornithine carbamyltransferase (OTC), and carbamate kinase (CK), catalyze the breakdown of arginine to ornithine, CO2, and two molecules of ammonia, with the production of ATP from ADP. The genes of the AD system, which are subject to complex regulation in the oral streptococci, have been isolated in bacteriophage lambda by screening for AD activity. The AD gene, designated arcA, was expressed from recombinant bacteriophage or in cells harboring plasmid subclones from this phage at a level up to 1,000-fold lower than the level in fully derepressed S. sanguis but apparently under the control of its own promoter. By subcloning in Escherichia coli mutants defective in anabolic OTC (argF argL) and CK (carB), it was demonstrated that the genes for S. sanguis OTC and CK were located adjacent to the AD gene. The levels of expression of the OTC and CK genes (arcB and arcC, respectively) were also very low in E. coli, although arcC expression was not as poor as arcA and arcB expression when compared with the levels found in S. sanguis. Also, arcB and arcC were unable to complement the defects in their anabolic counterparts. Introduction of the entire AD system or subclones which encoded only the AD gene into E. coli harboring defects in arginine and pyrimidine biosynthesis resulted in a 10- to 15-fold decrease in the level of AD activity, suggesting that arginine or its metabolites may regulate AD expression. Transposon mutagenesis was utilized to construct defined mutants of S. sanguis with mutations in the AD gene cluster. AD gene expression in these mutants indicated that the expression of the AD genes in this organism is strongly interrelated. The isolation and partial characterization of the arc genes represents the first step in the genetic manipulation of the AD system in the oral streptococci for analysis of the regulation of AD, analysis of the role of the system in plaque ecology, and utilization of the system to modulate the cariogenicity of dental plaque.
常见的口腔细菌血链球菌可通过精氨酸脱亚氨酶（AD）系统降解精氨酸。该系统的三种酶，即AD、鸟氨酸氨甲酰基转移酶（OTC）和氨基甲酸激酶（CK），催化精氨酸分解为鸟氨酸、二氧化碳和两分子氨，并由ADP生成ATP。AD系统的基因在口腔链球菌中受到复杂调控，通过筛选AD活性已在λ噬菌体中分离得到。指定为arcA的AD基因，可从重组噬菌体中表达，或在含有该噬菌体质粒亚克隆的细胞中表达，其表达水平比完全去阻遏的血链球菌低1000倍，但显然受自身启动子控制。通过在合成代谢OTC（argF argL）和CK（carB）有缺陷的大肠杆菌突变体中进行亚克隆，证明血链球菌OTC和CK的基因位于AD基因附近。OTC和CK基因（分别为arcB和arcC）在大肠杆菌中的表达水平也非常低，尽管与血链球菌中的水平相比，arcC的表达不像arcA和arcB那么差。此外，arcB和arcC无法弥补其合成代谢对应物的缺陷。将完整的AD系统或仅编码AD基因的亚克隆引入精氨酸和嘧啶生物合成有缺陷的大肠杆菌中，导致AD活性水平降低10至15倍，这表明精氨酸或其代谢产物可能调节AD的表达。利用转座子诱变构建了血链球菌AD基因簇中有突变的特定突变体。这些突变体中AD基因的表达表明该生物体中AD基因的表达密切相关。arc基因的分离和部分表征是对口腔链球菌中AD系统进行基因操作的第一步，用于分析AD的调控、该系统在菌斑生态学中的作用，以及利用该系统调节牙菌斑的致龋性。
