The mechanism for N 2 activation in the E 4 - state of nitrogenase.

Nitrogenases take nitrogen from the air and reduce it to ammonia. It has long been known that N 2 becomes activated after four reductions in the catalytic cycle, in the E 4 state. Several mechanisms for the activation have been suggested. In the present study a previous mechanism has been revised based on recent experimental findings. In the present mechanism N 2 H 2 is formed in E 4 . As in the previously suggested mechanism, there are four initial reductions before catalysis (the A-states), after which a sulfide is released and the first state in catalysis (E 0 ) is formed. In E 4 , N 2 becomes bound and protonated in the Fe1, Fe2, Fe4 region, in which the hydrides have left two electrons. The rate-limiting step is the formation of N 2 H by a hydrogen atom transfer from Cys275 to N 2 bound to Fe4, concerted with an additional electron transfer from the cofactor. The mechanism fulfills all requirements set by experiments. The activation of N 2 is preceded by a formation of H 2 from two hydrides, the carbide is kinetically hindered from being protonated, the E 4 state is reversible. An important aspect is the presence of a water molecule in the Fe2, Fe6 region. The non-allowed formations of H 2 from a hydride and a proton have been investigated and found to have higher barriers than the allowed formation of H 2 from two hydrides.