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Journal article
Using Directed Evolution to Improve Hydrogen Production in Chimeric Hydrogenases from Algal Species
Enzyme and microbial technology, Vol.173, 110349
02/2024
PMID: 27702473
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Abstract
Algae generate hydrogen from sunlight and water utilizing high-energy electrons generated during photosynthesis. The amount of hydrogen produced in heterologous expression of the wild-type hydrogenase is currently insufficient for industrial applications. One approach to improve hydrogen yields is through directed evolution of the DNA of the native hydrogenase. Here, we created 113 chimeric algal hydrogenase gene variants derived from combining segments of three parent hydrogenases, two from Chlamydomonas reinhardtii (CrHydA1 and CrHydA2) and one from Scenedesmus obliquus (HydA1). To generate chimeras, there were seven segments into which each of the parent hydrogenase genes was divided and recombined in a variety of combinations. The chimeric and parental hydrogenase sequences were cloned for heterologous expression in Escherichia coli, and 40 of the resultant enzymes expressed were assayed for H2 production. Chimeric clones that resulted in equal or greater production obtained with the cloned CrHydA1 parent hydrogenase were those comprised of CrHydA1 sequence in segments #1, 2, 3, and/or 4. These best-performing chimeras all contained one common region, segment #2, the part of the sequence known to contain important amino acids involved in proton transfer or hydrogen cluster coordination. The amino acid sequence distances among all chimeric clones to that of the CrHydA1 parent were determined, and the relationship between sequence distances and experimentally-derived H2 production was evaluated. An additional model determined the correlation between electrostatic potential energy surface area ratios and H2 production. The model yielded several algal mutants with predicted hydrogen productions in a range of two to three times that of the wild-type hydrogenase. The mutant data and the model can now be used to predict which specific mutant sequences may result in even higher hydrogen yields. Overall, results provide more precise details in planning future directed evolution to functionally improve algal hydrogenases.
•Chimeric algal hydrogenase gene mutants were created from three genes of two microalgal species
•Swapping gene segments resulted in some constructs with improved hydrogen production
•Sequence and electrostatic potential models may help predict improved function with mutation
Details
- Title
- Using Directed Evolution to Improve Hydrogen Production in Chimeric Hydrogenases from Algal Species
- Publication Details
- Enzyme and microbial technology, Vol.173, 110349
- Resource Type
- Journal article
- Publisher
- Elsevier Inc; United States
- Format
- pdf
- Copyright
- © 2023 The Author(s). Published by Elsevier Inc. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/4.0/)
- Identifiers
- 99380478097306600
- Academic Unit
- Biology; Center for Environmental Diagnostics and Bioremediation ; Hal Marcus College of Science and Engineering
- Language
- English