Ortholog Analysis and Transformation of Glycoside Hydrolase Genes in Hyperthermophilic Archaeal Thermococcus Species

TL;DR

This study shows that integrating a gene cluster from one archaeal species into another can enhance its ability to break down complex sugars and produce hydrogen at high temperatures.

Contribution

The study demonstrates stable integration of a large GH gene cluster in hyperthermophilic archaea for enhanced biomass conversion.

Findings

T. onnurineus GH03 showed significantly higher growth when cultivated with laminarin.

The GH03 mutant produced more hydrogen compared to the wild-type strain.

Large-fragment DNA integration is feasible in hyperthermophilic archaea.

Abstract

Archaea thrive in extreme environments, exhibiting unique traits with significant biotechnological potential. In this study, we investigated whether Thermococcus onnurineus NA1 could stably integrate a large glycoside hydrolase (GH) gene cluster from T. pacificus P-4, enhancing β-linked polysaccharides degradation for hydrogen production. Among 35 Thermococcus genomes examined via OrthoFinder2 and OrthoVenn3, and selecting Tpa-GH gene clusters as the target, we cloned and integrated Tpa-GH into T. onnurineus NA1 using a fosmid-based system, creating the GH03 mutant. Cultivation in a modified MM1 medium supplemented with laminarin revealed significantly higher growth and hydrogen production in T. onnurineus GH03 than in the wild-type strain. Our findings demonstrate the feasibility of stable, large-fragment DNA integration in hyperthermophilic archaea and underscore the promise of T. onnurineus GH03 as a strain for high-temperature biomass conversion.

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