Distal Heme Pocket Residues of B-type Dye-decolorizing Peroxidase
Distal Heme Pocket Residues of B-type Dye-decolorizing Peroxidase
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B型染料脱色过氧化物酶远端血红素口袋残基
DOI：
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发表时间：
2012
影响因子：
4.8
通讯作者：
L. Eltis
中科院分区：
文献类型：
--
作者：
Rahul Singh;J. C. Grigg;Z. Armstrong;Michael E. P. Murphy;L. Eltis
背景：DypB是一种Dyp-type过氧化物酶，氧化Mn（II）并转化木质素。结果：DypB形成稳定的化合物I，其在D153 A和N246 A中迅速衰变为化合物II，但在R244 L变体中不可检测。结论：需要Arg-244而不是Asp-153来形成化合物I表明DyPs调节过氧化循环不同于植物过氧化物酶。意义：了解DyPs有助于利用其生物技术潜力。来自霍氏红球菌RHA 1的DypB是一种细菌染料脱色过氧化物酶（DyP），其氧化木质素和Mn（II）。三个残基相互作用的铁结合的溶剂物种在铁DypB：Asn-246和保守的Asp-153和Arg-244。用丙氨酸取代Asp-153或Asn-246对化合物I形成的二级速率常数（k1 ≤ 105 m-1 s-1）和H2 O2的特异性常数（kcat/Km）的影响最小。即使在D153 A/N246 A双变体中，这些值也降低了不到30倍。然而，与野生型酶（540 s）相比，这些取代显著降低了化合物I的稳定性（t1/2 ≤ 0.13 s）。相比之下，取代精氨酸-244与亮氨酸废除过氧化物酶的活性，和血红素铁的变体显示了pH值依赖性的过渡，从高自旋（pH 5）到低自旋（pH 8.5）。设计了两种变体来模拟植物过氧化物酶活性位点：D153 H，其与H2 O2的反应性降低了一个数量级以上，而N246 H，其没有检测到过氧化物酶活性。X射线晶体学研究表明，在变体的结构变化仅限于远端血红素环境。数据确立了Arg-244在DypB中化合物I形成中的重要作用，可能通过电荷稳定和质子转移。Asp-153和Asn-246的主要作用似乎是调节化合物I的后续反应性。这些结果扩大了已知在血红素过氧化物酶中催化化合物I形成的残基的范围。
Background: DypB, a Dyp-type peroxidase, oxidizes Mn(II) and transforms lignin. Results: DypB forms a stable Compound I that rapidly decays to Compound II in the D153A and N246A but is undetectable in the R244L variant. Conclusion: The requirement of Arg-244 but not Asp-153 to form Compound I indicates that DyPs modulate the peroxidative cycle differently than plant peroxidase. Significance: Understanding DyPs helps harness their biotechnological potential. DypB from Rhodococcus jostii RHA1 is a bacterial dye-decolorizing peroxidase (DyP) that oxidizes lignin and Mn(II). Three residues interact with the iron-bound solvent species in ferric DypB: Asn-246 and the conserved Asp-153 and Arg-244. Substitution of either Asp-153 or Asn-246 with alanine minimally affected the second order rate constant for Compound I formation (k1 ∼ 105 m−1s−1) and the specificity constant (kcat/Km) for H2O2. Even in the D153A/N246A double variant, these values were reduced less than 30-fold. However, these substitutions dramatically reduced the stability of Compound I (t½ ∼ 0.13 s) as compared with the wild-type enzyme (540 s). By contrast, substitution of Arg-244 with leucine abolished the peroxidase activity, and heme iron of the variant showed a pH-dependent transition from high spin (pH 5) to low spin (pH 8.5). Two variants were designed to mimic the plant peroxidase active site: D153H, which was more than an order of magnitude less reactive with H2O2, and N246H, which had no detectable peroxidase activity. X-ray crystallographic studies revealed that structural changes in the variants are confined to the distal heme environment. The data establish an essential role for Arg-244 in Compound I formation in DypB, possibly through charge stabilization and proton transfer. The principle roles of Asp-153 and Asn-246 appear to be in modulating the subsequent reactivity of Compound I. These results expand the range of residues known to catalyze Compound I formation in heme peroxidases.
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影响因子：
2.9
作者：
Ahmad, Mark;Roberts, Joseph N.;Bugg, Timothy D. H.
通讯作者：
Bugg, Timothy D. H.
影响因子：
2.9
作者：
L. Vitello;J. Erman;Mark A Miller;Jimin Wang;J. Kraut
通讯作者：
L. Vitello;J. Erman;Mark A Miller;Jimin Wang;J. Kraut
影响因子：
5.6
作者：
Goblirsch B;Kurker RC;Streit BR;Wilmot CM;DuBois JL
通讯作者：
DuBois JL
精氨酸 82 在细菌视紫红质光循环过程中快速质子释放中的作用：对含有 15N 标记精氨酸的紫色膜进行时间分辨 FT-IR 研究。
DOI：
10.1021/bi049238g
发表时间：
2004
期刊：
Biochemistry.
影响因子：
--
作者：
Xiao,Yaowu;Hutson,MShane;Belenky,Marina;Herzfeld,Judith;Braiman,MarkS
通讯作者：
Braiman,MarkS
影响因子：
2.9
作者：
Roberts, Joseph N.;Singh, Rahul;Eltis, Lindsay D.
通讯作者：
Eltis, Lindsay D.