Mitochondrial adenosinetriphosphatase inhibitor protein: reversible interaction with complex V (ATP synthetase complex)

Abstract

Mitochondrial ATPase inhibitor protein (IF1) reacts reversibly with complex V and inhibits up to 90% of its ATPase activity. Both the rate and extent of inhibition are pH and temperature dependent and increase as the pH is lowered from pH 8 tp 6.7 (the lowest pH examined) or as the temperature is increased from 4 to 36 degrees C. Nucleotide triphosphates plus Mg2+ ions are required for inhibition of complex V ATPase activity by IF1. In the presence of Mg2+ ions, the effectiveness order of nucleotides is ATP greater than ITP greater than GTP greater than UTP. Highly purified complex V, which requires added phospholipids for expressing ATPase and ATP-Pi exchange activities, cannot be inhibited by IF1 plust ATP-Mg2+ unless phospholipids are also added. This indicates that the active state of the enzyme is necessary for the IF1 effect to be manifested, because F1-ATPase, which does not contain nor require phospholipids for catalyzing ATP hydrolysis, can be inhibited by IF1 plus ATP-Mg2+ in the absence of added phospholipids. The IF1-inhibited complex V, but not IF1-inhibited F1-ATPase, can be reactivated by incubation at pH greater than 7.0 in the absence of ATP-Mg2+. The reactivation rate is pH dependent and is influenced by temperature and enzyme concentration. Complex V preparations contain small and variable amounts of IF1. This endogenous IF1 behaves the same as added IF1 with respect to conditions described above for inhibition and reactivation and can result in 25-50% inhibition in different complex V preparations. However, complex V lacking endogenous IF1 can be reconstituted from F0, F1, oligomycin sensitivity conferring protein, and phospholipids. Inhibition of this reconstituted preparation in the presence of ATP-Mg2+ depends entirely on addition of IF1. In general, the ATP-Pi exchange activity of complex V is more sensitive to the chemical inhibitors of F1-AtPase tha its ATPase activity. This is not so, however, for IF1. Under conditions that IF1 caused approximately 75% inhibition of ATPase activity of complex V, no more than 10% of the ATP-Pi exchange activity was inhibited.

Release of the inhibitory action of the natural ATPase inhibitor protein on the mitochondrial ATPase

“…8B). The values for the activation energy E, = 53.34 i 6.7 kJ/mol and the activation enthalpy AH* = 50.83 & 6.7 kJ/mol ( n = 7) obtained are similar to those reported by Gomez-Fernandez and Harris [26] and Galante et al [28] for the reversal of inhibition by AIP. However, the activation entropy we obtained A$* = 158 18.78 J/mol K, is higher than other reported values.…”

Membrane Topography and Near-neighbor Relationships of the Mitochondrial ATP Synthase Subunits e, f, and g

“…The latter has been reported to undergo a ␤-strand to ␣-helix conformation change as the medium pH is changed from 6.5 to 8.0 (28). This conformation change has functional significance because at pH Ͻ7.0, the inhibitor protein binds to F 1 in the presence of MgATP and inhibits ATP hydrolysis, and at pH Ͼ7.0, it is released (29). Whether e undergoes functionally significant conformation changes with pH also remains to be seen.…”

IF1, a natural inhibitor of mitochondrial ATP synthase, is not essential for the normal growth and breeding of mice

“…It is a nuclear-coded mitochondrial protein and is evolutionarily conserved from yeast to mammals [2,3]. IF1 forms a dimer at acidic pH (~6.7) and exhibits inhibitory effects, but a tetramer form, which is formed at basic pH (~8.0), cannot interact with ATP synthase [4–6]. The crystal structure of the F 1 -ATPase with bound IF1 shows that the C-terminal α-helix of IF1 is inserted into the interface between the α- and β-subunits of F 1 -ATPase [7].…”