Details of experiments between the fragments P57105~6-103 and P03126~149-158
Note that the results of all experiments are listed, regardless of the modification states of the fragments.
Experiment series 1
Protein A protein: SYNJ2BP
Protein A fragment: 6-103
Protein A site: PDZ
Protein A construct: his6-MBP-TEVsite-PDZ
Protein B protein: HPV16-E6
Protein B fragment: 149-158
Protein B site: PBM
Protein B modification: (TPO)156
Protein B sequence: SSRTRRETQL
Protein B construct: Biotin-ttds-SSRTRREpTQL
Average holdup BI: -0.05
Immobilized partner concentration in holdup experiment (10-6M): 18
Experiment method: CALIP_holdup
Details of affinity fitting: BSA and lysozyme internal standards + bacterial cell lysate + normal layout
PUBMED ID of publication: 36115835
Number of measurements: 1
The affinity was below the detection threshold of the assay.
Experiment series 2
Protein A protein: SYNJ2BP
Protein A fragment: 6-103
Protein A site: PDZ
Protein A construct: his6-MBP-TEVsite-PDZ
Protein B protein: HPV16-E6
Protein B fragment: 149-158
Protein B site: PBM
Protein B sequence: SSRTRRETQL
Protein B construct: biotin-ttds-SSRTRRETQL
Average holdup BI: 0.65
Holdup BI standard deviation: 0.2
Immobilized partner concentration in holdup experiment (10-6M): 15
Experiment method: CALIP_holdup
Details of affinity fitting: BSA and lysozyme internal standards + bacterial cell lysate + normal layout
PUBMED ID of publication: 36115835
Number of measurements: 8
Measured pKd: 5.18
Experiment series 3
Protein A protein: SYNJ2BP
Protein A fragment: 6-103
Protein A site: PDZ
Protein A construct: his6-MBP-TEVsite-PDZ
Protein B protein: HPV16-E6
Protein B fragment: 149-158
Protein B site: PBM
Protein B sequence: SSRTRRETQL
Protein B construct: biotin-ttds-SSRTRRETQL
Measured dissociation constant (10-6M): 3.5
Standard deviation of dissociation constant (10-6M): 0.08
Experiment method: competitive FP
Details of affinity fitting: Competitive binding equation in ProFit
PUBMED ID of publication: 36115835
Number of measurements: 3
Measured pKd: 5.46
Experiment series 4
Protein A protein: SYNJ2BP
Protein A fragment: 6-103
Protein A site: PDZ
Protein A construct: his6-MBP-TEVsite-PDZ
Protein B protein: HPV16-E6
Protein B fragment: 149-158
Protein B site: PBM
Protein B modification: (TPO)156
Protein B sequence: SSRTRRETQL
Protein B construct: Biotin-ttds-SSRTRREpTQL
Measured dissociation constant (10-6M): 0
Experiment method: competitive FP
Details of affinity fitting: Competitive binding equation in ProFit
PUBMED ID of publication: 36115835
Number of measurements: 3
The affinity was below the detection threshold of the assay.
Experiment series 5
Protein A protein: SYNJ2BP
Protein A fragment: 6-103
Protein A site: PDZ
Protein A construct: his6-MBP-TEVsite-PDZ
Protein B protein: HPV16-E6
Protein B fragment: 149-158
Protein B site: PBM
Protein B modification: (TPO)156
Protein B sequence: SSRTRRETQL
Protein B construct: Biotin-ttds-SSRTRREpTQL
Average holdup BI: -0.02
Normalized holdup BI: -0.02
Immobilized partner concentration in holdup experiment (10-6M): 18
Experiment method: DAPF_holdup
Details of affinity fitting: fluorescein + mCherry internal standards + normalization based on CALIP_holdup data + reverse layout
PUBMED ID of publication: 36115835
Number of measurements: 1
The affinity was below the detection threshold of the assay.
Experiment series 6
Protein A protein: SYNJ2BP
Protein A fragment: 6-103
Protein A site: PDZ
Protein A construct: his6-MBP-TEVsite-PDZ
Protein B protein: HPV16-E6
Protein B fragment: 149-158
Protein B site: PBM
Protein B sequence: SSRTRRETQL
Protein B construct: biotin-ttds-SSRTRRETQL
Average holdup BI: 0.58
Holdup BI standard deviation: 0.07
Normalized holdup BI: 0.7
Normalized holdup BI standard deviation: 0.08
Immobilized partner concentration in holdup experiment (10-6M): 18
Experiment method: DAPF_holdup
Details of affinity fitting: fluorescein + mCherry internal standards + normalization based on CALIP_holdup data + reverse layout
PUBMED ID of publication: 36115835
Number of measurements: 2
Measured pKd: 5.18
Experiment series 7
Protein A protein: SYNJ2BP
Protein A fragment: 6-103
Protein A site: PDZ
Protein A construct: his6-MBP-TEVsite-PDZ
Protein B protein: HPV16-E6
Protein B fragment: 149-158
Protein B site: PBM
Protein B sequence: SSRTRRETQL
Protein B construct: biotin-ttds-SSRTRRETQL
Average holdup BI: 0.51
Normalized holdup BI: 0.64
Immobilized partner concentration in holdup experiment (10-6M): 18
Experiment method: DAPF_holdup
Details of affinity fitting: fluorescein + mCherry internal standards + normalization based on CALIP_holdup data + reverse layout
Experimental details: not reported internal standards of holdup layout #5
Number of measurements: 1
Measured pKd: 5.06
Experiment series 8
Protein A protein: SYNJ2BP
Protein A fragment: 6-103
Protein A site: PDZ
Protein A construct: his6-MBP-TEVsite-PDZ
Protein B protein: HPV16-E6
Protein B fragment: 149-158
Protein B site: PBM
Protein B sequence: SSRTRRETQL
Protein B construct: biotin-ttds-SSRTRRETQL
Average holdup BI: 0.54
Normalized holdup BI: 0.65
Immobilized partner concentration in holdup experiment (10-6M): 18
Experiment method: DAPF_holdup
Details of affinity fitting: fluorescein + mCherry internal standards + normalization based on CALIP_holdup data + reverse layout
Experimental details: not reported internal standards of holdup layout #5
Number of measurements: 1
Measured pKd: 5.08
Experiment series 9
Protein A protein: SYNJ2BP
Protein A fragment: 6-103
Protein A site: PDZ
Protein A construct: his6-MBP-TEVsite-PDZ
Protein B protein: HPV16-E6
Protein B fragment: 149-158
Protein B site: PBM
Protein B sequence: SSRTRRETQL
Protein B construct: Biotin-ado-ado-SSRTRRETQL
Average holdup BI: 0.69
Normalized holdup BI: 0.72
Immobilized partner concentration in holdup experiment (10-6M): 18
Experiment method: DAPF_holdup
Details of affinity fitting: fluorescein + mCherry internal standards + normalization based on CALIP_holdup data + reverse layout
Experimental details: Crude peptide synthesis product. The peptide concentration in the affinity conversion step may differ from reality. Not reported internal standards of holdup layout #6
PUBMED ID of publication: 36115835
Number of measurements: 1
Measured pKd: 5.24
Experiment series 10
Protein A protein: SYNJ2BP
Protein A fragment: 6-103
Protein A site: PDZ
Protein A construct: his6-MBP-TEVsite-PDZ
Protein B protein: HPV16-E6
Protein B fragment: 149-158
Protein B site: PBM
Protein B sequence: SSRTRRETQL
Protein B construct: Biotin-ttds-SSRTRRETQL
Measured dissociation constant (10-6M): 33
Standard deviation of dissociation constant (10-6M): 4
Experiment method: SPR
PUBMED ID of publication: 30726710
Number of measurements: 1
Measured pKd: 4.48