Katsumata K, Hayakawa M, Tanaka M, Sugiyama S, Ozawa T
Department of Biomedical Chemistry, Faculty of Medicine, University of Nagoya, Japan.
Biochem Biophys Res Commun. 1994 Jul 15;202(1):102-10. doi: 10.1006/bbrc.1994.1899.
Point mutations, oxygen damage and deletions in the heart mitochondrial (mt) DNA of a 19-year-old male patient with premature aging, who died of mitochondrial cardiomyopathy, were comprehensively analyzed. With total base-sequencing, one syn- mutation in the tRNA(Asp) gene and one mit-mutation in the ND3 gene were demonstrated. Using microHPLC/MS, 0.20% of the total deoxyguanosine (dG) were proved to be converted into its hydroxy-radical adduct, 8-hydroxy-dG, of which amount corresponds to that in normal subjects of 78 years old. The total detection system for mtDNA deletions, using 180 kinds of primer pairs, revealed extensive fragmentation of mtDNA; 235 types of deletions existed with various sizes, 97 of which yielded mtDNA minicircles lacking both of the replication origins of light- and heavy-strands. Deleted mtDNA accounted for 84% of the total mtDNA. In a man died from an accident at age 28 having almost the same mtDNA genotype except syn-, 50 types of deleted mtDNA, accounting for 15% of the total, were detected in his heart mtDNA. These results will present a clue to an unidentified mechanism of somatic mtDNA replication and the molecular basis of aging heart.
对一名死于线粒体心肌病的19岁早衰男性患者心脏线粒体（mt）DNA中的点突变、氧化损伤和缺失进行了全面分析。通过全碱基测序，在tRNA（Asp）基因中发现了一个同义突变，在ND3基因中发现了一个线粒体突变。使用微HPLC/MS分析，发现总脱氧鸟苷（dG）中有0.20%转化为其羟基自由基加合物8-羟基-dG，其含量与78岁正常受试者相当。使用180种引物对的mtDNA缺失检测系统显示mtDNA存在广泛片段化；存在235种不同大小的缺失类型，其中97种产生了缺乏轻链和重链复制起点的mtDNA小环。缺失的mtDNA占总mtDNA的84%。在一名28岁死于事故且mtDNA基因型除同义突变外几乎相同的男性中，在其心脏mtDNA中检测到50种缺失的mtDNA，占总数的15%。这些结果将为尚未明确的体细胞mtDNA复制机制和衰老心脏的分子基础提供线索。
