PI (H+) AND HOMOLOGOUS MITOCHONDRIAL ANION TRANSPORTERS

PI (H ) 和同源线粒体阴离子转运蛋白

• 批准号: 3282982
• 负责人: Hartmut none Wohlrab
• 金额: $ 26.01万
• 依托单位: BOSTON BIOMEDICAL RESEARCH INSTITUTE
• 依托单位国家: 美国
• 财政年份: 1984
• 资助国家: 美国
• 起止时间: 1984-04-01 至 1992-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/3282982
• 关键词: Saccharomyces; acidity /alkalinity; adenine nucleotides; affinity labeling; chemical structure function; complementary DNA; gel electrophoresis; genetic manipulation; heart; ion transport; laboratory rabbit; laboratory rat; liver; membrane permeability; mitochondria; mitochondrial membrane; molecular cloning; nuclear magnetic resonance spectroscopy; phosphates; point mutation; protein reconstitution; protein sequence; protein structure; radiotracer; transport proteins

The mitochondrial phosphate transport protein (PTP) is responsible for the transport of most of the inorganic phosphate (Pi) across the inner mitochondrial membrane. This transport (Phosphate-Hydrogen cotransport) is essential for steady state oxidative phosphorylation as it is carried out by mitochondria in the cell: ADP is phosphorylated to ATP in the mitochondrial matrix while most of the ATP is utilized in the extra mitochondrial space, liberating Pi, which must be transported back into the mitochondria. We have identified this protein, reconstituted the transport activity of the highly purified protein, sequenced its 47 N-terminal amino acids, and have purified a bovine cardiac PTP cDNA clone. We have discovered significant sequence homologies between the mitochondrial ADP/ATP carrier and PTP. We plan to obtain the complete cDNA-derived amino acid sequence of PTP from bovine cardiac tissue and rat liver and the PTP gene from the yeast Saccharomyces cerevisiae. Homologous regions are expected to reflect evolutionarily conserved and catalytically essential segments of the protein. We have identified a photoaffinity label that acts specifically on the mitochondrial matrix exposed active site of PTP. We plan to identify the protein segment it reacts with. Other reagents will be used to probe the active site(s) and the topology of PTP in the mitochondrial membrane. We plan to use site-specific mutagenesis in Saccharomyces cerevisiae to identify catalytically essential amino acid residues or sequence segments of PTP. We also plan to purify and sequence the mammalian PTP gene, in order to identify tissue-specific control sequences and, together with the ADP/ATP carrier gene, determine whether there is a mitochondrial transport protein multigene family with members having similar control sequences, i.e. conserved sequence segments in the 5'-flanking, 5'-untranslated, intervening, and 3'-untranslated regions.

线粒体磷酸运输蛋白（PTP）负责