BDB -BiopanningDataSet Details

Biopanning Data

BiopanningDataSet ID: 3484

Peptides:

Sequences	Affinity Values
YYVSWPPDMMHY(11/17) GRTPLLDSFLAG(3/17) TQQDYAWLLDH(1/17) SYSNFPLLHSYW(1/17) HDTWPLLATKTS(1/17)	[500 ± 25 nM, 299.3 nM] [~43 μM, NA] [NA] [NA] [NA]

Sequences Features:

Motif:

[PL]LL[DAH]

Unique Sequence Number:

Experimental Method:

Phage display (common panning)

Sequencing Platform:

Sequencing Method:

Sanger sequencing

Rounds of Panning:

Reference:

PMID: 32659860

Target Name:

Signal transducer and activator of transcription 3

Template Name:

Not determined.

Structure of Target-Template Complex:

Not determined.

Structure of Target-Peptide Complex:

Not determined.

Library Name:

Ph.D.-12 phage display library (X12)

Affinity Measurement Method:

ELISA

Affinity Measurement Description:

The affinity (Kd) of the peptides to STAT3 was determined by isothermal titration calorimetry (ITC) using no ITC calorimeter (TA Instruments, DE, USA). The alysis of which shows that peptide 2 interacted with higher affinity (Kd = 500 ± 25 nM) while peptide 1 was a slightly weaker (Kd = 1.79 ± 0.12 μM). Peptide 3, however, showed nearly 80x weaker compared to peptide 2 (Kd ~ 43 μM). All the three peptides were found to bind with a release of heat. For peptide ELISA, the strength of the association was recorded by Cytation 3 Imaging reader (BioTek Instruments). The result shows that the peptides interacted with STAT3 in a dose‐dependent manner. Peptide 1 was shown to have an affinity of 299.3 nM while the value for peptide 2 is 232.9 nM. Peptide 3 did not reach saturation under the identical conditions, suggesting a weaker interaction with STAT3. The first column of the affinity values is the Kd value determined by ITC. The second column is the strength of the association determined by peptide ELISA.

Brief Description:

A 100‐fold dilution of the PhD‐12™ library (New England Biolabs) was then added and incubated for 1 hr.The precipitated phages were pelleted by centrifugation (60 min, 16,100 g, 4°C), and the tubes dried for 20 min at room temperature (RT). The pellets were then re‐suspended in 1 ml of TBST buffer containing 0.02% w/v N3 and 1% (w/v) BSA. After re‐precipitation and suspension, the top 80% of the supertant was used for the next round of phage amplifications and selection.

BiopanningDataSet Comments:

The YYVSWPPDMMHY sequence was found to be enriched by 36% while another with a short consensus motif was displayed in 20% of the phages. Binding alysis by isothermal titration calorimetry shows the most displayed peptide interacted with a Kd of 1.79 μM, which on modification of its structure to mimic the tural binding partners of STAT3 brought the affinity to high nomolar range (Kd = 500 nM). Using a panel of tumor cell lines, we show that the peptides prevented the proliferation of triple‐negative breast cancer cells with a moderate activity (GI50 = 50 μM).

Curator:

Chunchao Pu

Curation Date:

2020-12-01

First |

Prev |

3484

| Next

| Last