BDB -BiopanningDataSet Details

Biopanning Data

BiopanningDataSet ID: 3365

Peptides:

Sequences

Affinity Values

GETRAPL(3)
TVRTSAD(2)
YPSPWGY(2)
ISLPSPT(1)
SSQLSRP(1)
YNSPTHH(1)
APKQSLE(1)
APTAVSK(1)
ETTHLTG(1)
GGLSSRP(1)
GVPTALP(1)
KPFPPMK(1)
LGTSMQL(1)
LPLWEVY(1)
LTAQPST(1)
MISSNSS(1)
QLKPLEF(1)
QLVYPAP(1)
SGTHHKA(1)
SLGPSPG(1)
SPHDVAYD(1)
TLNRVPN(1)
TNTMTRA(1)
TPHPLRL(1)
VNRSSLY(1)
VSTPSTP(1)

[n.a.]
[n.a.]
[n.a.]
[0.25 ± 0.02]
[0.24 ± 0.01]
[0.30 ± 0.03]
[n.a.]
[n.a.]
[n.a.]
[n.a.]
[n.a.]
[n.a.]
[n.a.]
[n.a.]
[n.a.]
[n.a.]
[n.a.]
[n.a.]
[n.a.]
[n.a.]
[n.a.]
[n.a.]
[n.a.]
[n.a.]
[n.a.]
[n.a.]

Sequences Features:

Motif:

Unique Sequence Number:

Experimental Method:

Phage display (common panning)

Sequencing Platform:

Sequencing Method:

Sanger sequencing

Rounds of Panning:

Reference:

PMID: 30268850

Target Name:

Cystic fibrosis (CF) mucus

Template Name:

Not determined.

Structure of Target-Template Complex:

Not determined.

Structure of Target-Peptide Complex:

Not determined.

Library Name:

Ph.D.-7 phage display library (X7)

Affinity Measurement Method:

ELISA

Affinity Measurement Description:

To understand binding affinities between phage-displayed peptides and the major protein component of mucus (mucin), ELISA was performed. A mucin solution was used instead of CF mucus since other mucus components such as lecithin or BSA might block the binding of mucin protein to the wells. Phage clones SSQ-phage, ISL-phage, and YNS-phage binding to mucin was measured by ELISA and compared to wild-type phage. The anti-M13 antibody used for detection binds specifically to an epitope on the pVIII major coat protein. Thus the binding affinities measured correlate with phage-mucin interactions. M13KE wild-type phage demonstrated a statistically significant higher binding affinity for mucin than phage clones SSQ-phage, ISL-phage, and YNS-phage Mucin only and monoclonal antibody only controls did not show specific binding, confirming that the measured binding affinities are specific to interactions between phage and mucin. The affinity value was measured at 450nm. The absorbance (450nm) value was reproduced from Figure 5 and shown.

Brief Description:

An initial concentration of 2e10 phage pfu was mixed with cystic fibrosis (CF) mucus, and 250 μL of the mixture was transferred to the apical compartment (donor) in the transwell system containing 1.5 mL phosphate buffered saline (PBS) in the basolateral compartment (receiver) and incubated for 1 h at room temperature (25°C). After 1 h, the eluate was collected from the basolateral side and titered using standard double-layer plaque assay to quantify phage concentration. The eluted phage library was amplified in XL-1 Blue E. coli (Agilent, CA) to make more copies, which were quantified by plaque assay prior to the next round of screening. Equivalent amounts of input phage were added for each round. This iterative screening was performed for a total of four rounds.

BiopanningDataSet Comments:

Here, phage clones displaying discovered peptides were shown to have up to 2.6-fold enhanced diffusivity in the CF mucus model. In addition, we demonstrate reduced binding affinities to mucin compared to wild-type control.

Curator:

Lixu Jiang

Curation Date:

2019-01-29

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