Details
| Original language | English |
|---|---|
| Pages (from-to) | 154-161 |
| Number of pages | 8 |
| Journal | Biotechnology and bioengineering |
| Volume | 91 |
| Issue number | 2 |
| Publication status | Published - 15 Jun 2005 |
| Externally published | Yes |
Abstract
A biosensor for fructose determination was used as basis of an assay for the determination of glucosyltransferase (GTF) activities and applied to monitoring recombinant enzyme production. GTFs catalyze the synthesis of glucans from sucrose leading to the release of fructose. Specific fructose determinations in the μM concentration range were achieved with a fructose electrode based on fructose dehydrogenase, which was immobilized on a screen-printed platinum electrode. This electrode was used as basis of the new assay for GTF activity determinations. Depending on the amount of enzyme, the assay was completed within 15-30 min compared to 1-2 h for the traditional photometric assay. From the amount of fructose released in a given reaction time, GTF activities were determined down to approx. 20 U/L. Even unpurified samples from a recombinant GTF-S production process could be analyzed without any problems, and a good correlation was obtained to data obtained from the photometric assay. Analysis of samples from cultures of various rGTF-S-producing recombinant E. constrains grown on different media with SDS-PAGE and with the new assay identified the same strain and culture medium as optimum for recombinant GTF-S production.
Keywords
- Amperometric detection, Dextransucrase, Enzyme activity, Fructose dehydrogenase, Glucosyltransferase, Recombinant GTF-S, Screen-printed electrode
ASJC Scopus subject areas
- Biochemistry, Genetics and Molecular Biology(all)
- Biotechnology
- Chemical Engineering(all)
- Bioengineering
- Immunology and Microbiology(all)
- Applied Microbiology and Biotechnology
Cite this
- Standard
- Harvard
- Apa
- Vancouver
- BibTeX
- RIS
In: Biotechnology and bioengineering, Vol. 91, No. 2, 15.06.2005, p. 154-161.
Research output: Contribution to journal › Article › Research › peer review
}
TY - JOUR
T1 - Development of a rapid, quantitative glucosyltransferase assay based on a screen-printed fructose enzyme electrode and application to optimization studies on gtfD expression in recombinant Escherichia coli
AU - Kadow, S.
AU - Betiku, E.
AU - Rinas, U.
AU - Bilitewski, U.
PY - 2005/6/15
Y1 - 2005/6/15
N2 - A biosensor for fructose determination was used as basis of an assay for the determination of glucosyltransferase (GTF) activities and applied to monitoring recombinant enzyme production. GTFs catalyze the synthesis of glucans from sucrose leading to the release of fructose. Specific fructose determinations in the μM concentration range were achieved with a fructose electrode based on fructose dehydrogenase, which was immobilized on a screen-printed platinum electrode. This electrode was used as basis of the new assay for GTF activity determinations. Depending on the amount of enzyme, the assay was completed within 15-30 min compared to 1-2 h for the traditional photometric assay. From the amount of fructose released in a given reaction time, GTF activities were determined down to approx. 20 U/L. Even unpurified samples from a recombinant GTF-S production process could be analyzed without any problems, and a good correlation was obtained to data obtained from the photometric assay. Analysis of samples from cultures of various rGTF-S-producing recombinant E. constrains grown on different media with SDS-PAGE and with the new assay identified the same strain and culture medium as optimum for recombinant GTF-S production.
AB - A biosensor for fructose determination was used as basis of an assay for the determination of glucosyltransferase (GTF) activities and applied to monitoring recombinant enzyme production. GTFs catalyze the synthesis of glucans from sucrose leading to the release of fructose. Specific fructose determinations in the μM concentration range were achieved with a fructose electrode based on fructose dehydrogenase, which was immobilized on a screen-printed platinum electrode. This electrode was used as basis of the new assay for GTF activity determinations. Depending on the amount of enzyme, the assay was completed within 15-30 min compared to 1-2 h for the traditional photometric assay. From the amount of fructose released in a given reaction time, GTF activities were determined down to approx. 20 U/L. Even unpurified samples from a recombinant GTF-S production process could be analyzed without any problems, and a good correlation was obtained to data obtained from the photometric assay. Analysis of samples from cultures of various rGTF-S-producing recombinant E. constrains grown on different media with SDS-PAGE and with the new assay identified the same strain and culture medium as optimum for recombinant GTF-S production.
KW - Amperometric detection
KW - Dextransucrase
KW - Enzyme activity
KW - Fructose dehydrogenase
KW - Glucosyltransferase
KW - Recombinant GTF-S
KW - Screen-printed electrode
UR - http://www.scopus.com/inward/record.url?scp=23844506852&partnerID=8YFLogxK
U2 - 10.1002/bit.20501
DO - 10.1002/bit.20501
M3 - Article
C2 - 15915514
AN - SCOPUS:23844506852
VL - 91
SP - 154
EP - 161
JO - Biotechnology and bioengineering
JF - Biotechnology and bioengineering
SN - 0006-3592
IS - 2
ER -