Part:BBa_K633003
Extracellular Succrase Sacc
The extracellular sucrase SacC from Zymomonas mobilis differs from SacB as it has sucrase but not levansucrase activity. A subcellular fractionation of Escherichia coli BL21 carrying the plasmid pLSS2811 showed that more than 70% of the sucrase activity could be detected in the cytoplasmic fraction, suggesting that the enzyme was soluble and not secreted. Its structure is monomeric, with an optimum temperature of 20 to 40 ° C, pH of 2.5-7.5 and a size of 48 kDa. It also has a free N-terminus. This approach was important in order to attach it to membrane protein ompA, which displays its N-terminus on E. Coli’s outer membrane.
Invertase hydrolyses the non-reducing β-d-fructofuranoside residues of sucrose to yield inverted sugar. Exploiting this property we can measure its activity as it can also react with 3.5 dinitrosalicylic acid (DNS). The color change produced is proportional to the amount of inverted sugar released, which in turn is proportional to the invertase activity present in the sample. The absorbance is measured at 540 nm and converted into micromoles of reducing sugar produced using a calibration curve.
One invertase unit is defined as the amount of enzyme which will produce 1 micromole of reducing sugar (expressed as inverted sugar) per minute under the conditions specified in the procedure.(Gurunathan & Gunasekaran, 2004)
This sac C gene we used was cloned from Zymmomonas mobilis genome in the FEMSA Biotechnology Center by phD. Jose Manuel Aguilar, in team TEC-Monterrey's campus.
References:
Kannan R, Mukundan G, Aït-Abdelkader N, Augier-Magro V, Baratti J, Gunasekaran P. Molecular cloning and characterization of the extracellular sucrase gene (sacC) of Zymomonas mobilis. Arch Microbiol. 1995 Mar;163(3):195-204.
Gurunathan, S., & Gunasekaran, P. (2004). Differential expression of Zymomonas mobilis sucrase genes (sacB and sacC) in Escherichia coli and sucrase mutants of Zymomonas mobilis. Brazilian Archives of Biology Technology, 47(3)
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25INCOMPATIBLE WITH RFC[25]Illegal AgeI site found at 907
- 1000INCOMPATIBLE WITH RFC[1000]Illegal BsaI.rc site found at 607
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