# Heterologous expression and characterization of polyhydroxyalkanoate synthase genes from haloarchaeal strains in Haloferax mediterranei

## TL;DR

Scientists studied how different genes from salt-loving microbes can produce a biodegradable plastic with better properties.

## Contribution

A systematic framework for evaluating haloarchaeal PHA synthase functionality in a heterologous host.

## Key findings

- Recombinant strains produced PHBV, showing host-dependent limitations in native strains.
- PhaEC from H. jeotgali produced high-molecular-weight PHBV with enhanced 3HV composition.
- Propionate addition improved 3HV content without significant toxicity in H. jeotgali-derived PHBV.

## Abstract

Polyhydroxyalkanoates (PHAs) are promising materials for building a sustainable society due to their excellent biodegradability. Some haloarchaea, which require high salt concentrations for growth, possess class III PHA synthases (PhaECs) and produce poly(3-hydroxybutyrate-co-3-hydroxyvalerate) (PHBV). Since PhaECs are a major factor in determining monomer compositions and molecular weights that affect the physical properties of PHAs, understanding their functionalities is essential to widen their applications. This study aimed to systematically evaluate the functionality of haloarchaeal phaECs under uniform conditions using a heterologous expression system in a PHA-negative mutant of Haloferax mediterranei.

To investigate the characteristics of phaECs from the five strains of haloarchaea, the genes were introduced into the strain of H. mediterranei lacking the original phaEC. The recombinant strains were subjected to PHA production evaluation.

All transformants produced PHBV, whereas some native strains did not, indicating host-dependent limitation. The molecular weights of PHBVs produced by the strains possessing phaECs from Halalkalicoccus jeotgali and Natronococcus occultus were higher than those of PHBVs produced by H. mediterranei. In the presence of propionate, a precursor of 3-hydroxyvalerate (3HV), a strain containing phaEC derived from H. jeotgali boosted the 3HV composition in PHBV without significantly suppressing cell growth or decreasing molecular weight due to propionate toxicity.

These results demonstrated that phaECs from haloarchaeal strains other than H. mediterranei also has the potential to produce high-molecular-weight-PHBV. Notably, phaEC from H. jeotgali exhibited exceptional potential for producing PHBV with both high molecular weight and enhanced 3HV composition. This work establishes a systematic framework for functional genomics of haloarchaeal PHA synthases and provides insight for engineering tailor-made bioplastics using extremophiles.