Biochemistry Online: An Approach Based on Chemical Logic

CHAPTER 8 - OXIDATION/PHOSPHORYLATION

E: NITROGENASE - A REDUCTIVE USE OF METAL CENTERS

BIOCHEMISTRY - DR. JAKUBOWSKI

04/15/16

Learning Goals/Objectives for Chapter 8D:

describe the structure, metal cofactors, and ligands of nitrogenase;
describe the path of electrons from the mobile electron carrier to the P cluster of nitrogenase;
discuss the role of ATP in the nitrogenase reaction;
draw and describe the Lowe and Thorneley cycle to show the sequential additions of electrons and protons to nitrogenase;
describe the properties of the E4 Janus intermediate and its role in backward and forward reactions in the cycle;
describe the role of H₂ in the mechanism of nitrogenase;
describe the organometallic reactions oxidative addition and reductive elimination and their role in nitrogenase;
explain the mechanisms and changes in oxidation states for Fe ions and substrates/products for the first and second half of the nitrogenase reaction.

E2. The Structure of Nitrogenase

Nitrogenase is a multiprotein complex containing the following:

Two identical subunits, E and F, which have binding sites for the mobile carrier of electrons (the protein ferrodoxin or flavodoxin), ATP, and an FeS cofactor (4Fe-4S, called the F cluster) which accept electrons. These subunits are hence called the nitrogenase reducatase subunits
a heterodimer of an alpha and beta subunits. The a (alpha chain) binds the 8Fe-7S F cluster and the Fe-S-Mo M cluster. These subunits comprise the (di)nitrogenase subunits

The overall structure of the protein complex with bound ATP and metal centers are shown below.

NitrogenaseTotalStructure

An interactive Jsmol structure of nitrogenase is found in the link below.

JSMol (HTML5)

Protopedia: Nitrogenase

An enhanced view of the bound cofactors and ATP are shown in the same spatial orientation in the figure below.

NitrogenasesWithATPMetalComplexes

The metal centers are shown below in more detail in both line and space fill views.

metalclustersNitrogenase

Mo is bound to 3 sulfur ions and two a OH and carboxyl group of 3-hydroxy-3-carboxy, adipic acid in the crystal structure of shown above.

The M cluster has an interstitial carbide ion that derives from -CH₃ attached to the sulfur of S-adenosyl-methionine (SAM) allowing the carbide to be labeled with either ¹³C or ¹⁴C for mechanistic studies. These labeled carbides are not exchanged or used as a substrate when the enzyme undergoes catalytic turnover. Hence it seems that the carbide probably just stabilizes the M cluster. it won't be shown in the figures below showing more detailed mechanisms.

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