Department of Biology, Georgia State University, Atlanta, GA 30303, USA.
J Bacteriol. 2010 Nov;192(22):5874-80. doi: 10.1128/JB.00673-10. Epub 2010 Sep 10.
In comparison to other pseudomonads, Pseudomonas aeruginosa grows poorly in L-lysine as a sole source of nutrient. In this study, the ldcA gene (lysine decarboxylase A; PA1818), previously identified as a member of the ArgR regulon of L-arginine metabolism, was found essential for L-lysine catabolism in this organism. LdcA was purified to homogeneity from a recombinant strain of Escherichia coli, and the results of enzyme characterization revealed that this pyridoxal-5-phosphate-dependent decarboxylase takes L-lysine, but not L-arginine, as a substrate. At an optimal pH of 8.5, cooperative substrate activation by L-lysine was depicted from kinetics studies, with calculated K(m) and V(max) values of 0.73 mM and 2.2 μmole/mg/min, respectively. Contrarily, the ldcA promoter was induced by exogenous L-arginine but not by L-lysine in the wild-type strain PAO1, and the binding of ArgR to this promoter region was demonstrated by electromobility shift assays. This peculiar arginine control on lysine utilization was also noted from uptake experiments in which incorporation of radioactively labeled L-lysine was enhanced in cells grown in the presence of L-arginine but not L-lysine. Rapid growth on L-lysine was detected in a mutant devoid of the main arginine catabolic pathway and with a higher basal level of the intracellular L-arginine pool and hence elevated ArgR-responsive regulons, including ldcA. Growth on L-lysine as a nitrogen source can also be enhanced when the aruH gene encoding an arginine/lysine:pyruvate transaminase was expressed constitutively from plasmids; however, no growth of the ldcA mutant on L-lysine suggests a minor role of this transaminase in L-lysine catabolism. In summary, this study reveals a tight connection of lysine catabolism to the arginine regulatory network, and the lack of lysine-responsive control on lysine uptake and decarboxylation provides an explanation of L-lysine as a poor nutrient for P. aeruginosa.
与其他假单胞菌相比，铜绿假单胞菌在 L-赖氨酸中生长不良，L-赖氨酸是其唯一的营养来源。在本研究中，ldcA 基因（赖氨酸脱羧酶 A；PA1818）先前被鉴定为 L-精氨酸代谢的 ArgR 调控子的成员，对于该生物体的 L-赖氨酸分解代谢是必需的。LdcA 从重组大肠杆菌菌株中被纯化为均相，酶特性的结果表明，这种依赖吡哆醛 5-磷酸的脱羧酶以 L-赖氨酸为底物，但不以 L-精氨酸为底物。在最佳 pH8.5 下，从动力学研究中描绘了 L-赖氨酸的协同底物激活，计算出的 K(m)和 V(max)值分别为 0.73 mM 和 2.2 μmole/mg/min。相反，在野生型 PAO1 菌株中，ldcA 启动子被外源 L-精氨酸诱导，但不受 L-赖氨酸诱导，并且通过电泳迁移率变动分析证明了 ArgR 与该启动子区域的结合。这种对赖氨酸利用的特殊精氨酸控制也从摄取实验中得到了注意，其中在存在 L-精氨酸但不存在 L-赖氨酸的情况下，放射性标记的 L-赖氨酸的掺入增强。在缺乏主要精氨酸分解代谢途径的突变体和细胞内 L-精氨酸池的基础水平较高且因此升高 ArgR 反应性调控子（包括 ldcA）的情况下，在 L-赖氨酸上快速生长被检测到。当编码精氨酸/赖氨酸：丙酮酸转氨酶的 aruH 基因从质粒上组成型表达时，也可以增强 L-赖氨酸作为氮源的生长；然而，ldcA 突变体在 L-赖氨酸上的生长表明该转氨酶在 L-赖氨酸分解代谢中的作用较小。总之，本研究揭示了赖氨酸分解代谢与精氨酸调控网络的紧密联系，并且缺乏赖氨酸响应控制对赖氨酸摄取和脱羧的解释了 L-赖氨酸对铜绿假单胞菌来说是一种较差的营养物质。
