Difference between revisions of "Part:BBa K3712011"

Revision as of 08:32, 21 October 2021

Flexible peptide

USAGE AND BIOLOGY

We use it when connecting ELP with other proteins, because it helps maintain the conformation of the two proteins

RESULTS

After culturing, induction and sterilization, the results of nickel column affinity chromatography show that both of Wild-type BLP-7-27 ELP and Optimized BLP-7-27 ELP elution have single peaks at 80 mM with good symmetry, which preliminarily indicates that the target protein is purified. Then we take the BCA method to determine the concentration (Table1) and SDS-PAGE to verify the existence of product.Flexible peptide linking ELP and BLP-7 works well, but linking ELP and TLR2 antagonist with flexible peptide will cause peptide fragmentation.

Figure 1.A) The elution peak height of Wild-type BLP-7-27 ELP and Optimized BLP-7-27 ELP and the running gel spectrum after purification. A) The elution peak height of Wild-type BLP-7-27 ELP is 1.41; B) The height of the Optimized BLP-7-27 ELP elution peak is 1.26; C) The gel running map, lane 1/2 is two-tube stream, lane 3 For Wild-type BLP-7-27 ELP, a thicker band appears near the 15kDa marker; lane 4/5/6 is a three-pipe stream that collects Optimized BLP-7-27 ELP, and a thicker band appears near the 15kDa marker.

Future direction:

(1) Unfortunately, due to time issues, we have no way to complete the purification of TLR2 antagonist. In the future, we will try different linkers to connect Wild-type/Optimized TLR2 antagonist and 27ELP. (2) Although our purified samples will not affect the subsequent functional verification, it can be seen from SDS-PAGE that even the purified bands will also have miscellaneous bands. Therefore, we want to use molecular sieves and freeze dryers for further purification attempts to obtain purer samples and make our experiment more successful.

Sequence and Features