Ancient nitrogenases are ATP dependent

Area of Science:

• Biochemistry
• Evolutionary Biology
• Enzymology

Background:

• Life relies on energy currencies like adenosine triphosphate (ATP) for cellular processes.
• The evolutionary origins and ancient enzyme dependence on ATP remain unclear.
• Reconstructing ancestral enzymes offers insights into early biochemistry.

Purpose of the Study:

• To investigate the evolutionary necessity of ATP for ancient enzymes.
• To experimentally reconstruct a Proterozoic-era nitrogenase ancestor.
• To determine the nucleotide and metal ion specificity of the ancestral enzyme.

Main Methods:

• Experimental reconstruction of an ancestral nitrogenase enzyme.
• Assays to measure enzyme activity with various nucleotide triphosphates and divalent metal ions.
• Comparison of ancestral enzyme activity and efficiency with the extant enzyme.

Main Results:

• The reconstructed ancestral nitrogenase strictly required ATP, showing no activity with GTP, ITP, or UTP.
• Magnesium (Mg2+) was the preferred divalent metal ion, though others supported reduced activity.
• The ancestral enzyme exhibited an ATP hydrolysis efficiency of two molecules per electron transferred, matching the extant enzyme.

Conclusions:

• Provides direct experimental evidence for ATP usage by an ancient enzyme.
• Demonstrates a conserved, strict requirement for ATP in nitrogenase function across evolutionary time.
• Highlights the early establishment of ATP as a critical energy currency in biological systems.