Part:BBa_K523001
RBS + malS (E. coli periplasmic α-amylase)
This is the E. coli amylase gene malS. The part contains the native Ribosome Binding Site.
The part was made using the strategy outlined in BBa_K523000, and therefore contains 4 extra bases at the 5' end which generate a BglII restriction site.
Usage and Biology
The product protein is believed to be periplasmic and thus may not actually be capable of degrading starch, unless it leaks from the periplasm in significant quantities. Its natural function in E. coli presumably involves degrading shorter glucose chains.
The SignalP program predicts that a 17 amino acid localisation signal (at the N terminal) is cleaved off before the protein reaches its mature form.
Experiments
We (Edinburgh 2011) placed this part under the control of the lac promoter and streaked colonies on a starch agar plate. A negative control without this construct was also streaked out. The cells were incubated for 3 days.
One colony failed to grow for some reason, but the others did. We flooded the plate with iodine, which turns black in the presence of starch. The area under the negative control turned black, while the areas under the malS streaks did not. The iodine gradually evaporated, turning the plate clear again. After 30 minutes, a halo was seen around the two malS streaks, suggesting some starch degradation around them.
Before the assay. Colony 1 failed to grow.
Immediately after iodine flooding.
30 minutes after iodine flooding.
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12INCOMPATIBLE WITH RFC[12]Illegal NheI site found at 1233
- 21COMPATIBLE WITH RFC[21]
- 23COMPATIBLE WITH RFC[23]
- 25INCOMPATIBLE WITH RFC[25]Illegal AgeI site found at 369
Illegal AgeI site found at 1534 - 1000COMPATIBLE WITH RFC[1000]
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