New Enzyme Discovered in Drug-Resistant Bacteria

Researchers have identified the first known pyridoxal phosphate (PLP)-dependent enzyme involved in a chemical modification linked to bacterial responses to metabolic stress. The discovery of amino valeramididine synthetase (AvaS) provides new insight into how bacteria use RNA modifications to regulate protein production and adapt to changing conditions, including exposure to antibiotics. The work, published in Nature Chemical Biology, offers a potential new avenue for identifying targets for future antimicrobial treatments.

Understanding the new enzyme

AvaS is the enzyme responsible for producing a tRNA modification known as amino valeramide cytidine (ava²C) in Pseudomonas aeruginosa, a bacterium associated with serious infections including pneumonia and sepsis. Ava²C had previously been detected in several bacteria and plants, but the enzyme responsible for producing it was unknown. Using a high-throughput liquid chromatography-tandem mass spectrometry (LC-MS/MS) platform for RNA modification profiling, the team screened thousands of Pseudomonas aeruginosa mutants and identified AvaS. The researchers also confirmed the presence of ava²C in Acinetobacter baumannii and Vibrio cholerae, as well as in the plant Arabidopsis thaliana.

The study found that AvaS uses PLP, a derivative of vitamin B6, to convert the known modification lysidine (k²C) into ava²C. This is the first reported example of a PLP-dependent enzyme directly involved in tRNA modification. PLP-dependent enzymes have traditionally been associated with amino acid metabolism and related biochemical processes. The discovery places PLP-dependent enzymes in a previously unrecognized class of tRNA-modifying enzymes, adding a new chemical mechanism to those already known to regulate protein production, including methylation, thiolation and isomerization. It also identifies a new biological function for PLP-dependent enzymes, showing that they can directly modify tRNA as well as participate in metabolic processes.

The researchers found that ava²C affects how bacteria read genetic codes, allowing them to produce proteins more rapidly and efficiently and adapt to metabolic and oxidative stress. Dr Jingjing Sun, Research Scientist at SMART AMR, first author and co-corresponding author of the paper, said the finding shows for the first time that PLP-dependent enzymes can directly modify tRNA, opening new areas of research into RNA chemistry and bacterial adaptation. Prof Peter Dedon, Co-lead Principal Investigator at SMART AMR, Professor of Biological Engineering at MIT and co-corresponding author of the paper, said the discovery adds to growing knowledge of the functions of RNA modifications and could provide new insight into processes relevant to antimicrobial resistance and infectious disease.

Future research directions

The SMART AMR team plans to investigate how ava²C affects bacterial stress responses and metabolism, as well as whether disrupting the modification could affect bacterial survival. Since ava²C was also found in plants, future research could examine whether similar mechanisms occur in other organisms and how the modification affects protein production. The work also demonstrates the potential of SMART AMR’s epitranscriptomics platform for identifying previously unknown RNA-modifying enzymes on a large scale. The researchers said this could have applications in biotechnology and drug discovery, including the search for new targets for antimicrobial treatments.

The research was conducted at SMART with funding from Singapore’s National Research Foundation under its Campus for Research Excellence and Technological Enterprise (CREATE) program.