INFLUENCE OF ADP, AMP-PNP AND OF DEPLETION OF NUCLEOTIDES ON THE STRUCTURAL-PROPERTIES OF F(1)ATPASE - A FOURIER-TRANSFORM INFRARED SPECTROSCOPIC STUDY

Mitochondrial F(1)ATPase from beef heart mas treated with different bu ffers in order to modulate the nucleotide content of the enzyme and th en analysed by FT-IR spectroscopy, Treatment of F(1)ATPase with a buff er lacking nucleotides and glycerol led to the formation of two fracti ons consisting of an inactive aggregated enzyme deprived almost comple tely of bound nucleotides and of an active enzyme containing ATP only in the tight sites and having a structure largely accessible to the so lvent and a low thermal stability. Treatment of F(1)ATPase with satura ting ADP, which induced the hysteretic inhibition during turnover, or AMP-PNP did not affect remarkably the secondary structure of the enzym e complex but significantly increased its compactness and thermal stab ility, It was hypothesised that the formation of the inactive aggregat ed enzyme was mainly due to the destabilisation of the alpha-subunits of F(1)ATPase and that the induction of the hysteretic inhibition is r elated to a particular conformation of the enzyme, which during turnov er becomes unable to sustain catalysis.