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Artificial microRNA-mediated knockdown of pyruvate formate lyase (PFL1) provides evidence for an active 3-hydroxybutyrate production pathway in the green alga Chlamydomonas reinhardtii

Paper ID Volume ID Publish Year Pages File Format Full-Text
23651 43460 2012 10 PDF Available
Title
Artificial microRNA-mediated knockdown of pyruvate formate lyase (PFL1) provides evidence for an active 3-hydroxybutyrate production pathway in the green alga Chlamydomonas reinhardtii
Abstract

Artificial microRNA technology was investigated as a means of down regulating metabolic pathways in the green alga Chlamydomonas reinhardtii, targeting pyruvate formate lyase (PFL1), which catalyzes the conversion of pyruvate to acetyl-CoA and formate during anoxic conditions. Two transformants with an 80–90% reduction in target protein and mRNA levels were identified. Nuclear magnetic resonance spectroscopy confirmed a substantial decrease in the production of formate in the knockdown lines during dark anoxic conditions and a re-routing of metabolism leading to enhanced production of ethanol and lactate. Under microaerobic conditions in the light, induced by sulphur-deprivation, knock-down of PFL1 resulted in reduced formate and ethanol production, increased net consumption of acetate and the excretion of lactate but no increase in the production of hydrogen. In addition the production of 3-hydroxybutyrate was identified in knock-down line cultures during the transition between microaerobic and anoxic conditions. Overall our results indicate that microRNA knock-down is a useful tool to manipulate anaerobic metabolism in C. reinhardtii.

► We investigate re-directing metabolic flux to increase H2 production in Chlamydomonas reinhardtii. ► Artificial microRNA technology was used to knock down pyruvate formate lyase. ► PFL1 knockdown caused reduced formate and ethanol production. ► PFL1 knockdown did not result in increased H2 production in the light during sulphur depletion. ► PFL1 knockdown resulted in the production of 3-hydroxybutyrate.

Keywords
3-HB, 3-hydroxybutyrate; ACK, acetate kinase; ADH1, bifunctional acetaldehyde/alcohol dehydrogenase; ALDH, aldehyde dehydrogenase; amiRNA, artificial microRNA; HP, sodium hypophosphite; HYDA1, [FeFe]-hydrogenase; LDH, d-lactate dehydrogenase; PAT, phospha
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Artificial microRNA-mediated knockdown of pyruvate formate lyase (PFL1) provides evidence for an active 3-hydroxybutyrate production pathway in the green alga Chlamydomonas reinhardtii
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Publisher
Database: Elsevier - ScienceDirect
Journal: Journal of Biotechnology - Volume 162, Issue 1, 30 November 2012, Pages 57–66
Authors
, , , , ,
Subjects
Physical Sciences and Engineering Chemical Engineering Bioengineering
Get Full-Text Now
Don't Miss Today's Special Offer
Price was $35.95
You save - $31
Price after discount Only $4.95
100% Money Back Guarantee
Full-text PDF Download
Online Support
Any Questions? feel free to contact us