Berlutti Francesca, Passariello Claudio, Selan Laura, Thaller Maria Cristina, Rossolini Gian Maria
Dipartimento di Scienze di Sanità Pubblica, Università di Roma 'La Sapienza', Piazzale A. Moro, 5, 00185 Rome, Italy1.
Dipartimento di Biologia, Università di Roma 'Tor Vergata', 00133 Rome, Italy2.
Microbiology (Reading). 2001 Oct;147(Pt 10):2831-2839. doi: 10.1099/00221287-147-10-2831.
Chryseobacterium meningosepticum is an aerobic Gram-negative rod widely distributed in natural environments. Unlike many bacteria, it produces a phosphate-irrepressible periplasmic alkaline phosphatase (AP). This work describes cloning of the gene encoding that enzyme from C. meningosepticum CCUG 4310 (NCTC 10585), and preliminary characterization of its product. The gene, named pafA, encodes a protein (PafA) of 546 amino acids with a calculated molecular mass of the mature peptide of 58682 Da. PafA exhibits high sequence identity with the PhoV AP of Synechococcus PCC 7942 (49.9% identity) and with the Cda Ca(2+)-dependent ATPase of Myroides odoratus (51.9% identity), while being more distantly related to the PhoD AP of Zymomonas mobilis (22.1% identity) and to the PhoA AP of Escherichia coli (14.0% identity). PafA was partially purified; it exhibits optimal activity at pH 8.5 and is active towards a broad spectrum of substrates including both phosphomonoesters and ATP, with preferential activity for the latter compound. The present findings allow definition of a new family of APs including 60 kDa, periplasmic enzymes whose expression is not influenced by freely available P(i) in the medium. Moreover, PafA can be considered an evolutionary intermediate between Ca(2+)-ATPase of M. odoratus and the APs PhoV of Synechococcus PCC 7942 and PhoD of Z. mobilis.
脑膜败血金黄杆菌是一种需氧革兰氏阴性杆菌，广泛分布于自然环境中。与许多细菌不同，它能产生一种不受磷酸盐抑制的周质碱性磷酸酶（AP）。这项工作描述了从脑膜败血金黄杆菌CCUG 4310（NCTC 10585）中克隆编码该酶的基因，并对其产物进行了初步表征。该基因名为pafA，编码一种由546个氨基酸组成的蛋白质（PafA），成熟肽的计算分子量为58682 Da。PafA与聚球藻PCC 7942的PhoV AP（同一性为49.9%）以及气味类芽孢杆菌的Cda Ca(2+)依赖性ATP酶（同一性为51.9%）具有高度序列同一性，而与运动发酵单胞菌的PhoD AP（同一性为22.1%）和大肠杆菌的PhoA AP（同一性为14.0%）的亲缘关系较远。PafA被部分纯化；它在pH 8.5时表现出最佳活性，对包括磷酸单酯和ATP在内的多种底物都有活性，对后一种化合物具有优先活性。目前的研究结果确定了一个新的AP家族，包括60 kDa的周质酶，其表达不受培养基中游离无机磷（P(i)）的影响。此外，PafA可被视为气味类芽孢杆菌的Ca(2+) - ATP酶与聚球藻PCC 7942的PhoV AP和运动发酵单胞菌的PhoD AP之间的进化中间体。
