Kennaway N G, Stankova L, Wirtz M K, Weleber R G
Department of Medical Genetics, Oregon Health Sciences University, Portland 97201.
Am J Hum Genet. 1989 Mar;44(3):344-52.
The purpose of this study was to characterize the mutant enzyme in nine patients with gyrate atrophy of the choroid and retina associated with ornithine aminotransferase (OAT) deficiency, to elucidate the mechanism of response to pyridoxine in four pyridoxine-responsive patients, and to determine the extent of genetic heterogeneity in both groups of patients. We have measured the apparent Km for pyridoxal phosphate (PLP) in fibroblast mitochondria and the heat stability of OAT at 45 degrees C in the presence and absence of PLP, using a sensitive radiochemical assay. The apparent Km for PLP was higher in pyridoxine-responsive patients than in nonresponsive patients whose apparent Km for PLP was normal. In contrast, the apparent Km for ornithine was normal in the seven patients studied. Surprisingly, the responsive patient with mildest clinical disease had the highest Km for PLP. However, she had the most stable enzyme, which presumably contributed to her milder phenotype. Western blot analyses of mitochondrial proteins, using antibody to human OAT, indicated clearly detectable OAT protein in pyridoxine-responsive patients and in two of five nonresponders, but low or undetectable levels in the other three patients. These data clarify the mechanism of pyridoxine response and indicate heterogeneity within as well as between the pyridoxine-responsive and the nonresponsive patients with gyrate atrophy.
本研究的目的是对9例患有与鸟氨酸转氨酶（OAT）缺乏相关的脉络膜和视网膜回旋状萎缩的患者体内的突变酶进行表征，阐明4例对吡哆醇有反应的患者对吡哆醇的反应机制，并确定两组患者的基因异质性程度。我们使用灵敏的放射化学分析法，测定了成纤维细胞线粒体中磷酸吡哆醛（PLP）的表观Km值，以及在有和没有PLP存在的情况下，OAT在45摄氏度时的热稳定性。对吡哆醇有反应的患者中PLP的表观Km值高于对吡哆醇无反应的患者，后者的PLP表观Km值正常。相比之下，所研究的7例患者中鸟氨酸的表观Km值正常。令人惊讶的是，临床症状最轻的有反应患者的PLP Km值最高。然而，她的酶最稳定，这可能是导致其较轻表型的原因。使用抗人OAT抗体对线粒体蛋白进行的蛋白质印迹分析表明，在对吡哆醇有反应的患者以及5例无反应者中的2例中可清楚检测到OAT蛋白，但在其他3例患者中水平较低或无法检测到。这些数据阐明了吡哆醇反应的机制，并表明在对吡哆醇有反应和无反应的回旋状萎缩患者内部以及之间均存在异质性。
