H2S HEMOGLOBIN I STRUCTURE--SITE DIRECTED MUTAGENESIS STUDY

血红蛋白I从Lucina pectinata可能是唯一的血红素蛋白，履行其功能，在硫化氢结合铁状态和双氧结合亚铁状态。一种提出的机制表明，Hbl与H2S反应形成血红素硫化铁低自旋复合物，将H2S运输到细菌共生体。硫化氢被认为是释放到细菌形成的亚铁蛋白质的电子转移从一个仍然未知的还原剂。此外，假定血红素-Fe/+3 SH2部分和GInE7之间存在H-键合以稳定HbISH2中心。同样，PheB10和PheE11残基分别与HbICO、HbIO2和HBICN中心之间存在极性相互作用。该假设是Hbl可能具有两种不同的配体稳定机制：一种用于Hbl-SH 2中心，另一种用于HbICO、HB10 2和HbICN部分。本文提出的工作旨在揭示附近氨基酸与HbISH2、HbIO2、HbICO和HbICN部分之间的相互作用。将使用定点诱变方法。在E7、B10和E11位置的Hbl定点变体的选择，以及与共振和FTIR振动光谱耦合的配体的同位素标记将阐明Hbl-配体双重稳定化机制和结构。我们计划解决的因素，控制H2S的结合，运输和交付的Hbl。现在，关于血红素蛋白中观察到的配体亲和力差异是否来自静电效应、空间约束或氢键差异，正在进行激烈的争论和讨论。从广义上讲，Hbl提供了解决这些问题的真实的优势，因为在这里，铁蛋白质以及亚铁蛋白质在其携带其生物学功能时是重要的、独特的和可逆的。本项目的一个长期目标是阐明好氧Lucina pectinata和未知厌氧细菌之间的共生相互作用和交流。

Hemoglobin I from Lucina pectinata is probably the only heme protein that carries out its function in the hydrogen-sulfide bound ferric state and in the dioxygen bound ferrous state. A proposed mechanism suggests that Hbl reacts with H2S to form a heme ferric sulfide low spin complex to transport the H2S to a bacteria symbion. Hydrogen sulfide is believed to be released to the bacteria upon formation of the ferrous protein by electron transfer from a still unknown reductant. Furthermore, the presence of H-bonding between the heme-Fe/+3 SH2 moiety and GInE7 is postulated to stabilize the HbISH2 center. Likewise, polar interactions between the PheB10 and PheE11 residues and the HbICO, HbIO2, and HBICN centers, respectively. This hypothesis is that Hbl may possesses two distinct ligand stabilization mechanism: one for the Hbl-SH2 center and the other for the HbICO, HBIO2, and HbICN moieties. The work proposed here is intended to unravel the interactions between the nearby amino acids and the HbISH2, HbIO2, HbICO, and HbICN moieties. A site-directed mutagenesis approach will be used. Choices of Hbl site-directed variants in the E7, B10, and E11 positions, and isotope labeling of ligands coupled to resonance and FTIR vibrational spectroscopies will clarify the Hbl-ligand dual stabilization mechanism and structure. We plan to address the factors that controls the binding, transport, and delivery of H2S by Hbl. Right now, there are intense arguments and discussions going on about whether the ligand affinity differences that are observed in heme proteins arise from electrostatic effects, from steric constraints, or from hydrogen-bonding differences. In the broad sense, Hbl offers a real advantage to address these issues as here the ferric, as well as the ferrous, protein is significant, unique and reversible when it carries its biological function. A long term goal of this project is to clarify the symbiotic interaction and communication between the aerobic Lucina pectinata and the unknown anaerobic bacteria.