Is Trehalose an Effective Quenching Agent of Azotobacter vinelandii Mo-Nitrogenase Turnover?

H₂-evolution assays, plus EPR and FTIR spectroscopies, using CO-inhibited Azotobacter vinelandii Mo-nitrogenase have shown that the disaccharide trehalose is an effective quenching agent of enzymatic turnover and also stabilizes the reaction intermediates formed. Complete inhibition of H₂-evolution activity was achieved at 1.5 M trehalose, which compares favorably to the requirement for 10 M ethylene glycol to achieve similar inhibition. Reaction-intermediate stabilization was demonstrated by monitoring the EPR spectrum of the 'hi-CO' form of CO-inhibited N₂ase, which did not change during 1 hr after trehalose quenching. Similarly, in situ photolysis with FTIR monitoring of 'hi-CO' resulted in the same 1973 and 1681 cm^-1 signals as observed previously in ethylene glycol-quenched systems. [a] These results clearly show that 1.5 M trehalose is an effective quench and stabilization agent for Mo-N₂ase studies. Possible applications are discussed.