Part:BBa_K1469002
UDP-GlcDH
UDP-glucose 6-dehydrogenase of Bacillus megaterium. The enzyme catalyses the NAD dependent oxidation of UDP-glucose to UDP-glucuronic acid, a key precursor molecule for hyaluronic acid production.
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000COMPATIBLE WITH RFC[1000]
Characterization by SSTi-SZG(2018)
This is an improvement of this part and gtaB (BBa_K1469005), which were originally introduced by Saarland iGEM in 2014. We characterized this part by co-overexpressing it with gtaB (BBa_K1469005) together in an operon. UDP-GlcDH is also known as tuaD in B.subtilis, and its gene product may contribute positively to HA production.
In our project, expression of the operon tuaD-gtaB was regulated under the control of a constitutive promoter P43, this operon is used for further increasing the production of the HA in B.subtilis. Gene products of tuaD and gtaB regulate the last two steps in the synthetic pathway of UDP-GlcUA, one of the two precursors of HA.
Fig1 the synthesis pathway of HA . In our experiment, by conducting CTAB experiments that form turbidity from a reaction between HA and CTAB solution, the results showed a remarkable increase in HA production when co-overexpressed tuaD-gtaB together (488mg/L, a 38% increase) (Figure2),In addition, Molecular weight analysis studies showed that HAs synthesized were high molecular weight(Figure3 ).
Figure 2: CTAB analysis of HA concentraton, a. Illustration of the turbidity by mixing different source of HA with CTAB solution. b: effects of overexpressing the precursor genes on HA production in recombinant B.
Figure 3: molecular weights of HA produced by overexpression of precursor genes in recombinant B. subtilis 168E strains using viscometer analysis.
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