Protein family shows how life has adapted to oxygen

Cornell scientists have created an evolutionary model that links organisms living in today’s oxygen-rich atmosphere to a time billions of years ago when Earth’s atmosphere was oxygen-deficient – by analyzing ribonucleotide reductases (RNRs), a family of proteins Used by all free living organisms and many viruses to repair and replicate DNA.

“By understanding the evolution of these proteins, we can understand how nature adapts to environmental changes at the molecular level. In turn, we also learn about our planet’s past,” said Nozomi Ando, ​​associate professor of chemistry and chemical biology at the College of Arts and Sciences and corresponding author of the study. “Comprehensive phylogenetic analysis of the ribonucleotide reductase family reveals an ancestral group,” published in eLife on October 4th.

Co-first authors of the study are Audrey Burnim and Da Xu, PhD students in chemistry and chemical biology, and Matthew Spence, Research School of Chemistry, Australian National University, Canberra. Colin J. Jackson, Professor of Chemistry, Australian National University, Canberra, is a corresponding author.

This undertaking included a large data set of 6,779 RNR sequences; Calculating the phylogeny took seven months (1.4 million CPU hours) on multiple high-performance computers combined. The approach, made possible by computational advances, opens up a new way to study other diverse protein families that have evolutionary or medical significance.

RNRs have adapted to changes in the environment over billions of years to maintain their catalytic mechanism due to their essential role in all DNA-based life, Ando said. Her lab studies protein allostery – how proteins are able to change their activity in response to the environment. The evolutionary information in a phylogeny gives us a way to study the relationship between a protein’s primary sequence and its three-dimensional structure, dynamics, and function.

RNRs are thought to be ancient in origin because they catalyze the reaction of converting RNA building blocks into DNA building blocks, Ando said, making them ideal for finding a molecular record.

“This chemistry would have been needed to transition from the hypothetical RNA world to the DNA/protein world that we currently live in,” Ando said. “It is also clear from the cofactors that RNRs use that this family of enzymes has adapted to the increase in oxygen in the Earth’s atmosphere. Both transitions happened billions of years ago.”

When scientists construct a phylogeny of a protein family, they calculate how the currently existing sequences arose, Ando said. In this process they have to estimate what happened in the past in order to get the sequences that exist now.

The researchers calculated RNR phylogeny by collecting a dataset of more than 100,000 sequences and curating it down to a computationally manageable dataset of 6,779 sequences while maintaining the diversity of the entire family, Burnim said. The sequences are approximately 400 to 1,100 amino acids in length. Using models of how amino acids mutate, they compared the sequences to see when they diverged.

From this work, the researchers discovered a new set of RNRs that would explain how two distinct adaptations to oxygen on Earth arose within this family of proteins.

They used small-angle X-ray scattering at the Cornell High Energy Synchrotron Source, cryogenic electron microscopy at the Cornell Center for Materials Research, and the AlphaFold2 artificial intelligence program to study the RNR of Synechococcus phage S-CBP4, a virus carrying a cyanobacterium, they said xu

“When we calculated the RNR pedigree, it turned out that there was a branch of RNRs that we didn’t know was a distinct lineage,” Ando said. “This branch included sequences from marine organisms, including cyanophages. Our characterization of one of the sequences suggests that there was an early adaptation to oxygen, and cyanobacteria are credited with supplying oxygen to the Earth.”

The results support the idea that molecular adjustments to oxygen occurred much earlier than the large-scale environmental changes on the planet as dated by the geochemical record, Ando said.

This first-ever unified evolutionary model for all classes of RNRs could show many future directions for the field, Xu said.

Ando plans to use the same approach to study how enzymes with the same overall structure evolved to catalyze vastly different chemical reactions.