MAMMALIAN ATP SYNTHASE: NOVEL INSIGHTS INTO THE ROLES OF ITS SUPERNUMERARY SUBUNITS

ATP synthase;Supernumerary Subunits;Biochemistry

Lee, Han GilPedersen, Peter L. (Pete) ;

Johns Hopkins University

关键词: ATP synthase;    Supernumerary Subunits;    Biochemistry;   

Others  :  https://jscholarship.library.jhu.edu/bitstream/handle/1774.2/58742/LEE-DISSERTATION-2015.pdf?sequence=1&isAllowed=y

瑞士|英语

来源: JOHNS HOPKINS DSpace Repository

【 摘 要 】

ATP synthase (FOF1) in mammals is an important and well recognized complex located on the inner mitochondrial membrane. It synthesizes ATP from ADP and Pi in the presence of Mg2+ by utilizing an electro-chemical gradient of protons across this membrane generated by an electron transport chain. The overall process is referred to as ;;oxidative phosphorylation”. More recent studies reported within or near the past decade have shown that the ATP synthase complex exists also on the plasma membrane of several mammalian tissues and have referred to it as the ;;ectopic” ATP synthase. At this location the ATP synthase may have important functions other than a role in ATP synthesis, or in addition to a role in ATP synthesis. Also, of some interest have been the findings that some of the soluble components of the ATP synthase such as subunit F6 and the regulator IF1 are present in human serum and interact with and affect the activity of the ectopic ATP synthase. For these reasons six soluble non catalytic subunits (d, OSCP, Factor B, F6 and IF1) of mitochondrial ATP synthase were investigated in some detail to determine to what extent they independently interact with the catalytic F1 component of the ATP synthase and influence its activity. Of particular interest were subunits d, OSCP, and F6, as these are known as ;;supernumerary subunits”, i.e., non-catalytic or extra subunits whose roles are not well defined.Maltose-Binding Protein (MBP) pull-down assays done in in a phosphate saline (PBS) buffer containing Triton X-100 showed that subunits d, F6, Factor B and OSCP can individually interact with the catalytic F1 subcomplex. These interactions were not affected appreciably by either conditions containing more salt (NaCl) or the addition of ATP, ADP and Mg2+, properties that may be common among mammalian species and also allowed within the mitochondrial structure. We also found that each subunit tested could independently affect the catalytic ATP hydrolytic activity by measuring the free phosphate released from ATP catalyzed by the isolated F1-ATPase in the presence of each tested subunit. Finally, we carried out an MBP pull-down assay without the detergent (Triton X-100) being present and reconfirmed these interactions. These data indicate that each of the soluble non-catalytic subunit components of ATP synthase in mammals can bind to the catalytic F1 subcomplex and affect its activity. Therefore, we suggests that within many animal and human cells such regulation of ATP synthase is likely to occur both at its well established mitochondrial location, and at its more recently established cell membrane location.