The biosynthesis of histidine, a proteinogenic amino acid, has been extensively studied due to its importance in bacterial growth and survival. Histidinol-phosphate phosphatase (Hol-Pase), which is responsible for the penultimate step of histidine biosynthesis, is generally the last enzyme to be characterized in many bacteria because its origin and evolution are more complex compared to other enzymes in histidine biosynthesis. However, none of the enzymes in histidine biosynthesis, including Hol-Pase, have been characterized in Pseudomonas aeruginosa, which is an important opportunistic gram-negative pathogen that can cause serious human infections. In our previous work, a transposon mutant of P. aeruginosa was found to display a growth defect on glucose-containing minimal solid medium. In this study, we found that the growth defect was due to incomplete histidine auxotrophy caused by PA0335 inactivation. Subsequently, PA0335 was shown to encode Hol-Pase, and its function and enzymatic activity were investigated using genetic and biochemical methods. In addition to PA0335, the roles of 12 other predicted genes involved in histidine biosynthesis in P. aeruginosa were examined. Among them, hisC2(PA3165), hisH2(PA3152), and hisF2(PA3151) were found to be dispensable for histidine synthesis, whereas hisG(PA4449), hisE(PA5067), hisF1(PA5140), hisB(PA5143), hisI(PA5066), hisC1(PA4447), and hisA(PA5141) were essential because deletion of each resulted in complete histidine auxotrophy; similar to PA0335, hisH1(PA5142) or hisD(PA4448) deletion caused incomplete histidine auxotrophy. Taken together, our results outline the histidine synthesis pathway of P. aeruginosa.

IMPORTANCE Histidine is a common amino acid in proteins. Because it plays critical roles in bacterial metabolism, its biosynthetic pathway in many bacteria has been elucidated. However, the pathway remains unclear in P. aeruginosa, an important opportunistic pathogen in clinical settings; in particular, there is scant knowledge about histidinol-phosphate phosphatase (Hol-Pase), which has a complex origin and evolution. In this study, P. aeruginosa Hol-Pase was identified and characterized. Furthermore, the roles of all other predicted genes involved in histidine biosynthesis were examined. Our results illustrate the histidine synthesis pathway of P. aeruginosa. The knowledge obtained from this study may help in developing strategies to control P. aeruginosa-related infections. In addition, some enzymes of the histidine synthesis pathway from P. aeruginosa might be used as elements of histidine synthetic biology in other industrial microorganisms.

由于组氨酸在细菌生长和存活中的重要性，因此已广泛研究了组氨酸（一种蛋白氨基酸）的生物合成。组氨酸生物合成的倒数第二个步骤中的组氨酸磷酸磷酸酶（Hol-Pase）通常是许多细菌中表征的最后一种酶，因为与组氨酸生物合成中的其他酶相比，其起源和进化更为复杂。但是，铜绿假单胞菌没有鉴定出组氨酸生物合成中的任何酶，包括Hol-Pase，铜绿假单胞菌是一种重要的机会革兰氏阴性病原体，可引起严重的人类感染。在我们以前的工作中，铜绿假单胞菌的转座子突变体发现在含葡萄糖的基本固体培养基上显示出生长缺陷。在这项研究中，我们发现生长缺陷是由于PA0335失活引起的组氨酸营养缺陷而导致的。随后，显示PA0335编码Hol-Pase，并使用遗传和生化方法研究其功能和酶活性。除PA0335以外，还检查了铜绿假单胞菌中其他12个与组氨酸生物合成有关的预测基因的作用。其中，hisC2（PA3165），hisH2（PA3152）和hisF2（PA3151）可用于组氨酸合成，而hisG（PA4449），hisE（PA5067），hisF1（PA5140），hisB（PA5143），hisI（PA5066），hisC1（PA4447）和hisA（PA5141）是必不可少的，因为它们各自的缺失会导致完全的组氨酸营养缺陷。与PA0335相似，hisH1（PA5142）或hisD（PA4448）缺失会导致组氨酸营养缺陷型。两者合计，我们的结果概述了铜绿假单胞菌的组氨酸合成途径。

重要信息组氨酸是蛋白质中的常见氨基酸。由于它在细菌代谢中起关键作用，因此已经阐明了其在许多细菌中的生物合成途径。然而，在临床环境中一种重要的机会病原体铜绿假单胞菌中，该途径尚不清楚。尤其是，关于组氨醇磷酸磷酸酶（Hol-Pase）的知识很少，它具有复杂的起源和发展。在这项研究中，鉴定并鉴定了铜绿假单胞菌Hol-Pase。此外，检查了所有其他预测的基因在组氨酸生物合成中的作用。我们的结果说明了铜绿假单胞菌的组氨酸合成途径。从这项研究中获得的知识可能有助于制定控制策略铜绿假单胞菌相关感染。另外，来自铜绿假单胞菌的组氨酸合成途径的一些酶可以用作其他工业微生物中组氨酸合成生物学的元素。