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A fast and sensitive coupled enzyme assay for the measurement of l-threonine and application to high-throughput screening of threonine-overproducing strains

Paper ID Volume ID Publish Year Pages File Format Full-Text
17111 42643 2014 7 PDF Available
Title
A fast and sensitive coupled enzyme assay for the measurement of l-threonine and application to high-throughput screening of threonine-overproducing strains
Abstract

•We develop a fast and sensitive coupled enzyme assay for the measurement of l-threonine (LTA).•LTA assay can distinguish the threonine concentration in 0.5 mM difference in some certain mediums.•LTA assay was successfully achieved to measure l-threonine in industrial fermentation samples.•LTA assay was applied to high throughput screening for threonine overproducing strains.•LTA assay is simple, accurate, specific and suitable for determination of threonine concentration in large numbers of samples.

Amino acids are important fermentation products that are used in the food, animal feed, pharmaceutical, and cosmetics industries. Development of a better quantitation assay to measure amino acid levels will facilitate research and application in medical and industrial fields. Here we report the development of an assay that allows the rapid measurement of threonine concentration. l-threonine aldolase (LTA) catalyzes the cleavage of threonine to glycine and acetaldehyde. The resulting acetaldehyde is further converted into ethanol by alcohol dehydrogenase accompanying NADH reduction. Therefore, monitoring the changes in NADH concentration can serve as a readout for the amount of threonine present within certain range. This LTA assay can distinguish a 0.5 mM difference in threonine concentration in some certain mediums. We successfully utilized this assay to measure l-threonine levels in industrial fermentation samples. Moreover, we were able to scale up this assay to 96-well format and use it in high-throughput screening of threonine-overproducing strains. We show this assay is simple, accurate, specific and suitable for determination of threonine concentration in large numbers of samples.

Keywords
l-Threonine aldolase; Threonine determination; Coupled enzyme assay; High throughput screening
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A fast and sensitive coupled enzyme assay for the measurement of l-threonine and application to high-throughput screening of threonine-overproducing strains
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Publisher
Database: Elsevier - ScienceDirect
Journal: Enzyme and Microbial Technology - Volume 67, December 2014, Pages 1–7
Authors
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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