STRUCTURE OF PROTOCATECHUATE 3,4-DIOXYGENASE FROM PSEUDOMONAS-AERUGINOSA AT 2.15 ANGSTROM RESOLUTION
STRUCTURE OF PROTOCATECHUATE 3,4-DIOXYGENASE FROM PSEUDOMONAS-AERUGINOSA AT 2.15 ANGSTROM RESOLUTION
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DOI：
10.1006/jmbi.1994.1754
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发表时间：
1994-12-16
影响因子：
5.6
通讯作者：
LIPSCOMB, JD
中科院分区：
文献类型：
--
作者：
OHLENDORF, DH;ORVILLE, AM;LIPSCOMB, JD
原儿茶酸3，4-双加氧酶通过掺入两个分子氧原子催化3，4-二羟基苯甲酸酯的芳环裂解，产生β-羧基-顺式，顺式-粘康酸。来自铜绿假单胞菌（现在重新分类为恶臭假单胞菌）的这种金属酶的结构已被细化到0.172至2.15埃分辨率的R因子。该结构是高度对称的（α β Fe 3+）（12）聚集体，具有均方根（r.m.s.）与原子相关的原子之间的差异为0.18埃。两种多肽（α和β）的三级结构是高度同源的（r.m.s.在127个C-α原子上相差1.05埃），表明祖先酶最初是具有两个活性位点的同源二聚体。事实上，非功能性残留活性位点保留了功能性活性位点的许多性质，但不结合铁。非血红素铁催化辅因子的配位几何结构可以最好地描述为三角双锥，其中Tyr 447（147 β）和His 462（162 β）用作轴向配体，Tyr 408（108 β）、His 460（160 β）和Wat 837用作赤道配体。活性位点环境具有许多碱性残基，其可促进酸性底物的结合。在位于α和β链之间的推定活性位点空腔内，五个近似共面的溶剂分子表明平面底物Trp 449（149 β），Ile 491（191 β）的位置，由Gly 14（14 α）和Pro 15（15 α）定义。在这个位置，Arg 457（157 β）的胍基将被底物掩埋，表明在催化中的功能作用。
Protocatechuate 3,4-dioxygenase catalyzes the aromatic ring cleavage of 3,4-dihydroxybenzoate by incorporating both atoms of molecular oxygen to yield beta-carboxy-cis, cis-muconate. The structure of this metalloenzyme from Pseudomonas aeruginosa (now reclassified as P. putida) has been refined to an R-factor of 0.172 to 2.15 Angstrom resolution. The structure is a highly symmetric (alpha beta Fe3+)(12) aggregate with a root-mean-square (r.m.s.) difference of 0.18 Angstrom among symmetry-related atoms. The tertiary structure of the two polypeptides (alpha and beta) are highly homologous (r.m.s. difference of 1.05 Angstrom over 127 C-alpha atoms), suggesting that the ancestral enzyme was originally a homodimer with two active sites. Indeed, a non-functional, vestigial active site retains many of the properties of the functional active site but does not bind iron. The coordination geometry of the non-heme iron catalytic cofactor can best be described as trigonal bipyramidal with Tyr447 (147 beta) and His462 (162 beta) serving as axial ligands, and Tyr408 (108 beta), His460 (160 beta) and Wat837 serving as equitorial ligands. The active site environment has a number of basic residues that may promote binding of the acidic substrate. Within the putative active site cavity which is located between alpha and beta chains, five approximately coplanar solvent molecules suggest a position for the planar substrate Trp449 (149 beta), Ile491 (191 beta), defined by Gly14 (14 alpha) and Pro15 (15 alpha). In this position the guanidino group of Arg457 (157 beta) would be buried by the substrate, suggesting a functional role in catalysis.