MECHANISMS OF IRON-A CQUISITION FROM HEMOGLOBIN IN PORPHYROMONAS GINGIVALIS

在这项研究中，我们研究了牙龈卟啉单胞菌从血红蛋白中获得血红素的机制。牙龈假单胞菌381的缺铁细胞在基础培养基加试验底物如血红蛋白、氯化血红素、转铁蛋白和各种无机铁化合物中培养。牙龈假单胞菌381对氯化血红素、转铁蛋白和血红蛋白的Ks值分别为3.85、4.91和0.0017 mm。然而，所测试的无机铁化合物不支持银杏叶霉菌的生长。这些发现表明，在铁有限的环境下，牙龈假单胞菌比其他含铁化合物更有效地利用血红蛋白作为铁源。然后，用[~3H]标记的血红蛋白测定人血红蛋白与牙龈假单胞菌细胞的结合。血红蛋白的结合迅速、可逆和特异地发生。Scatchard对结合数据的分析显示了线性曲线图，表明存在一个结合蛋白群体。表观解离常数Kd为1.0？B10.19×10^~(-6)M，每细胞结合位点数为3.2？未标记的人血红蛋白能抑制血红蛋白的结合，但不能抑制氯化血红素和原卟啉Ix的结合。血清白蛋白、转铁蛋白、乳铁蛋白、过氧化氢酶和细胞色素c对结合蛋白的结合有部分抑制作用。当生物体在氯化血红素限制的条件下生长时，血红蛋白的结合力大大增加。这些结果表明，牙龈假单胞菌细胞可能通过其表面的特定结合部位与人血红蛋白相互作用，这是获得铁的初步步骤。

In this study, we investigated on the mechanisms of the acquisition of heme from hemoglobin in Porphyromonas gingivalis. The iron-depleted cells of P.gingivalis 381 were incubated in the basal medium plus test substrates such as hemoglobin, hemin, transferrin and various inorganic iron-compounds. The value of Ks, a parameter analogous to Michaelis-Menten constant, was estimated.P.gingivalis 381 showed a Ks value of 3.85,4.91 and 0.0017 mM for hemin, transferrin and hemoglobin, respectively. However, the inorganic iron compounds tested did not support P.gingvalis growth. These findings suggest that P.gingivalis utilizes hemoglobin as an iron source much more effectively than other iron-containing compounds under an iron -limited environment. Next, binding of human hemoglobin by P.gingivalis cells was determined by using [^3H]-labeled hemoglobin. The binding of hemoglobin occurred rapidly, reversibly and specifically. Scatchard analysis of the binding data revealed linear plot, indicating the presence of one population of binding protein. The apparent Kd was 1.0? B10.19x10^<-6>M and number of binding sites per cell was 3.2? B10.76x10^4. Hemoglobin binding was inhibited by unlabeled human hemoglobin but not hemin and protoporphyrin IX.The binding was only partially inhibited by serum albumin, transferrin, lactoferrin, catalase and cytochrome c. These results suggest that the ligand recognized by the binding protein may not be the heme moiety. The binding of hemoglobin much increased when the organisms were grown in hemin-limited conditions. These results suggest that P.gingivalis cells may interact with human hemoglobin through specific binding sites on their surfaces as a preliminary step in iron acquisition.