Structure of PhnP, a Phosphodiesterase of the Carbon-Phosphorus Lyase Pathway for Phosphonate Degradation
Structure of PhnP, a Phosphodiesterase of the Carbon-Phosphorus Lyase Pathway for Phosphonate Degradation
复制标题
DOI：
10.1074/jbc.m808392200
复制
发表时间：
2009-06-19
影响因子：
4.8
通讯作者：
Jia, Zongchao
中科院分区：
文献类型：
--
作者：
Podzelinska, Kateryna;He, Shu-Mei;Jia, Zongchao
碳磷裂解酶是一种由phn操纵子编码的多酶系统，当无机磷缺乏时，它能使细菌代谢有机膦酸盐。通过序列同源性预测，由该操纵子编码的酶之一Phosphate是β-内酰胺酶超家族的金属依赖性水解酶。用各种水解敏感底物筛选表明，Phosphate是具有磷酸二酯酶活性的酶，对双（对硝基苯基）磷酸和2 '，3'-环核苷酸具有最大的特异性。未观察到对RNA的活性。金属离子依赖性的Phosphate与双（对硝基苯基）磷酸盐作为底物显示了一个明显的偏好Mn 2+和Ni 2+的催化，而Zn 2+提供穷人的活动。通过X射线晶体学以1.4埃分辨率解析Phosphorus的三维结构。Phosphorus的总体折叠与tRNase Z内切核酸酶的折叠非常相似，但缺乏这些酶用于结合其tRNA底物的长外位点模块。Phosphorus的活性位点可能包含两个Mn 2+离子，周围环绕着一系列活性位点残基，这些残基与tRNase Z酶中观察到的残基相同。第二，远程Zn 2+结合位点也被观察到，由一组半胱氨酸和组氨酸残基，严格保守的Phosphorus家族。该第二金属离子位点似乎稳定结构基序。
Carbon-phosphorus lyase is a multienzyme system encoded by the phn operon that enables bacteria to metabolize organophosphonates when the preferred nutrient, inorganic phosphate, is scarce. One of the enzymes encoded by this operon, PhnP, is predicted by sequence homology to be a metal-dependent hydrolase of the beta-lactamase superfamily. Screening with a wide array of hydrolytically sensitive substrates indicated that PhnP is an enzyme with phosphodiesterase activity, having the greatest specificity toward bis(p-nitrophenyl) phosphate and 2',3'-cyclic nucleotides. No activity was observed toward RNA. The metal ion dependence of PhnP with bis(p-nitrophenyl) phosphate as substrate revealed a distinct preference for Mn2+ and Ni2+ for catalysis, whereas Zn2+ afforded poor activity. The three-dimensional structure of PhnP was solved by x-ray crystallography to 1.4 angstrom resolution. The overall fold of PhnP is very similar to that of the tRNase Z endonucleases but lacks the long exosite module used by these enzymes to bind their tRNA substrates. The active site of PhnP contains what are probably two Mn2+ ions surrounded by an array of active site residues that are identical to those observed in the tRNase Z enzymes. A second, remote Zn2+ binding site is also observed, composed of a set of cysteine and histidine residues that are strictly conserved in the PhnP family. This second metal ion site appears to stabilize a structural motif.