ADP-RIBOSYLTRANSFERASE OF HELIOBACTER PYLORI

摘要幽门螺杆菌是慢性胃炎的病原。十二指肠溃疡。以及胃粘膜恶性肿瘤。探讨微生物发病机制的相关因素。我们研究了幽门螺杆菌是否产生adp -核糖基转移酶（ART），以及抗溃疡药物是否能抑制该酶的活性。对细菌进行超声培养，提取液与[腺苷酸- 32p] nadh孵育。当用sds-page和放射自显影分析检测混合物时，在细胞裂解液中观察到70kda蛋白（p70）以mg2 +依赖的方式进行放射性标记。p70的修饰似乎是由单一基因介导的，非酶和依赖性修饰的参与似乎不太可能。adp -核糖-P70连锁对羟胺处理敏感，arglnne和AGMATINE抑制P70的修饰，表明arglnne残基是P70的修饰位点。此外，胍丁氨酸可以被假定的酶以mg2 +依赖的方式用作底物。幽门螺杆菌ART不能被ARF激活，ARF是一种小的gtp结合蛋白，可刺激霍乱肠毒素的ART活性。一种抗溃疡的化合物，利巴米胺，可以抑制幽门螺杆菌细胞裂解物对p70和agmatine的adp -核糖基化。这表明酶可能是药物的靶标。这些结果表明幽门螺杆菌产生一种与细胞相关的arglnne特异性单art，它对抗溃疡药物敏感。此外。将幽门螺杆菌裂解物与HL-60细胞的单核生成物孵育，可引起一个50KDA蛋白的修饰，这表明该酶也可能对宿主细胞蛋白进行腺苷核苷化。由于许多细菌病原体产生的毒素具有单一活性，结果可能表明幽门螺杆菌在细菌的病理作用中起作用。

HELICOBACTER PYLORI IS AN ETIOLOGIC AGENT FOR CHRONlC GASTRITIS. DUODENAL ULCERS. AND GASTRIC MUCOUS MALIGNANCY. TO VESTIGATE FACTORS INVOLVED IN THE PATHOGENESIS OF THE MICROORGANISM. WE EXAMlNED WHETHER H. PYLORI PRODUCES AN ADP-RIBOSYLTRANSFERASE (ART) AND WHETHER AN ANTI-ULCER DRUG CAN INHIBIT THE ENZYME ACTIVITY. SONIFIED CULTURE OF THE BACTERIA WAS SUBJECTED TO ART ASSAY IN WHICH THE EXTRACT WAS INCUBATED WITH [ADENYLATE-32P]NAD. WHEN THE ASSAY MIXTURE WAS ANALYZED BY SDS-PAGE AND AUTORADIOGRAPHY, RADIOLABELELLING OF A 70KDA PROTEIN (P70) IN THE CELL LYSATE WAS OBSERVED IN A MG2+-DEPENDENT MANNER. THE MODIFICATION OF P70 SEEMED TO BE MEDIATED BY A MONO-ART, AND INVOLVEMENT OF NON-ENZYMATIC NAD-DEPENDENT MODIFICATIONS APPEARED TO BE UNLIKELY. THE OBSERVATIONS THAT (ADP-RIBOSE)-P70 LINKAGE WAS SUSCEPTIBLE TO HYDROXYLAMlNE TREATMENT AND THAT ARGlNlNE AND AGMATINE INHIBITED THE MODIFICATION OF P70 INDICATED THAT ARGlNlNE RESIDUE IN P70 IS THE SITE OF THE MODIFICATION. ADDITIONALLY, AGMATINE COULD BE UTILIZED AS A SUBSTRATE BY THE PUTATIVE ENZYME IN A MG2+-DEPENDENT MANNER. H. PYLORI ART WAS NOT ACTIVATED BY ARF, A SMALL GTP-BINDlNG PROTEIN WHICH IS STIMULATE ART ACTIVITY OF CHOLERA ENTEROTOXlN. AN ANTI-ULCER COMPOUND, REBAMIPIDE, INHIBITED THE ADP-RIBOSYLATION BOTH OF P70 AND OF AGMATINE BY H. PYLORI CELL LYSATE. SUGGESTlNG THAT THE ENZYME CAN BE A TARGET OF THE DRUG. THESE RESULTS INDICATE THAT H. PYLORI PRODUCES A CELL-ASSOCIATED ARGlNlNE-SPECIFIC MONO-ART, WHICH IS SUSCEPTIBLE TO ANTI-ULCER DRUG. FURTHERMORE. INCUBATION OF H. PYLORI LYSATE WITH THE MOMOGENATE OF HL-60 CELLS CAUSED THE MODIFICATION OF A 50KDA PROTElN, SUGGESTING THAT THE ENZYME MAY ALSO ADP-RIBOSYLATE HOST CELL PROTEIN. BECAUSE MANY BACTERIAL PATHOGEN PRODUCE TOXINS WITH MONO-ART ACTIVITY, THE RESULTS MAY SUGGEST THAT H. PYLORI ART PLAYS A ROLE IN PATHOLOGICAL EFFECT OF THE BACTERlUM.