黑色普氏菌与血红蛋白结合的特点及其血红蛋白结合蛋白的鉴定

Characterization of hemoglobin binding by Prevotella nigrescens and identification of hemoglobin binding protein

目的 :观察黑色普氏菌与血红蛋白结合的特点并鉴定血红蛋白结合蛋白。方法 :采用斑点印迹法观察黑色普氏菌的血红蛋白结合特点 ,利用SDS -PAGE及Westernblot法鉴定血红蛋白结合蛋白。结果 :热及胰蛋白酶预处理细菌使血红蛋白结合能力分别降低 6 8%和 35 % ,珠蛋白的抑制率为 81% ,乳铁蛋白部分抑制血红蛋白结合 ,转铁蛋白、细胞色素C和过氧化物酶对血红蛋白的结合皆无抑制作用。Westernblot结果显示 ,相对分子量Mr为 4.1× 10 4 ,5 .6× 10 4 和 5 .9× 10 4 的膜蛋白能与血红蛋白反应。结论 :4.1× 10 4 ,5 .6×10 4 和 5 .9× 10 4 的细胞膜蛋白可能参与了黑色普氏菌与血红蛋白的结合。

AIM: To characterize the hemoglobin binding activity by Prevotella nigrescens and to identify the hemoglobin binding protein. METHODS: Dot blot assay was adopted to characterize the hemoglobin binding activity.SDS-PAGE and Western blot were used to identify the hemoglobin binding protein. RESULTS: Heat and trypsin treatments of the bacteria decrease the hemoglobin binding activity by 68% and 35% respectively. The inhibition rate by globin was 81%. Lactoferrin also partially inhibited the activity, while transferrin, cytochrome C or catalase had little or no inhibitory effect. Although tested sugars did not affect the activity, some tested amino acids, such as arginine, cysteine, histidine and lysine, inhibited the activity partially. By Western blot assay, 4.1×10 4, 5.6 ×10 4 and 5.9×10 4 proteins of P. nigrescens reacted with hemoglobin. CONCLUSION: 4.1×10 4, 5.6 ×10 4 and 5.9×10 4membrane proteins of P. nigrescens may be involved in its hemoglobin binding.