Electron Transfer Precedes ATP Hydrolysis during Nitrogenase Catalysis

Document Type

Article

Language

eng

Format of Original

6 p.

Publication Date

10-2013

Publisher

National Academy of Sciences

Source Publication

Proceedings of the National Academy of Sciences

Source ISSN

0027-8424

Original Item ID

DOI: 10.1073/pnas.1311218110

Abstract

The biological reduction of N2 to NH3 catalyzed by Mo-dependent nitrogenase requires at least eight rounds of a complex cycle of events associated with ATP-driven electron transfer (ET) from the Fe protein to the catalytic MoFe protein, with each ET coupled to the hydrolysis of two ATP molecules. Although steps within this cycle have been studied for decades, the nature of the coupling between ATP hydrolysis and ET, in particular the order of ET and ATP hydrolysis, has been elusive. Here, we have measured first-order rate constants for each key step in the reaction sequence, including direct measurement of the ATP hydrolysis rate constant: kATP = 70 s−1, 25 °C. Comparison of the rate constants establishes that the reaction sequence involves four sequential steps: (i) conformationally gated ET (kET = 140 s−1, 25 °C), (ii) ATP hydrolysis (kATP = 70 s−1, 25 °C), (iii) Phosphate release (kPi = 16 s−1, 25 °C), and (iv) Fe protein dissociation from the MoFe protein (kdiss = 6 s−1, 25 °C). These findings allow completion of the thermodynamic cycle undergone by the Fe protein, showing that the energy of ATP binding and protein–protein association drive ET, with subsequent ATP hydrolysis and Pi release causing dissociation of the complex between the Feox(ADP)2 protein and the reduced MoFe protein.

Comments

Accepted version. Proceedings of the National Academy of Sciences, Vol. 110, No. 41 (October 2013): 16414-16419. DOI. © 2013 National Academy of Sciences. Used with permission.

Edwin Antony was affiliated with Utah State University at the time of publication.

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