Details of experiments between the fragments Q13424~84-171 and P27228~140-149
Note that the results of all experiments are listed, regardless of the modification states of the fragments.
Experiment series 1
Protein A protein: SNTA1
Protein A fragment: 84-171
Protein A site: PDZ
Protein A construct: his6-MBP-TEVsite-PDZ
Protein B protein: HPV35-E6
Protein B fragment: 140-149
Protein B site: PBM
Protein B sequence: SKPTRRETEV
Protein B construct: biotin-ttds-SKPTRRETEV
Average holdup BI: 0.69
Immobilized partner concentration in holdup experiment (10-6M): 39.7
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
Measured pKd: 4.79
Experiment series 2
Protein A protein: SNTA1
Protein A fragment: 84-171
Protein A site: PDZ
Protein A construct: his6-MBP-TEVsite-PDZ
Protein B protein: HPV35-E6
Protein B fragment: 140-149
Protein B site: PBM
Protein B sequence: SKPTRRETEV
Protein B construct: biotin-ttds-SKPTRRETEV
Measured dissociation constant (10-6M): 28.96
Standard deviation of dissociation constant (10-6M): 1.92
Experiment method: competitive FP
Details of affinity fitting: Competitive binding equation in ProFit
PUBMED ID of publication: 36115835
Number of measurements: 3
Measured pKd: 4.54
Experiment series 3
Protein A protein: SNTA1
Protein A fragment: 84-171
Protein A site: PDZ
Protein A construct: his6-MBP-TEVsite-PDZ
Protein B protein: HPV35-E6
Protein B fragment: 140-149
Protein B site: PBM
Protein B modification: (TPO)147
Protein B sequence: SKPTRRETEV
Protein B construct: Biotin-ttds-SKPTRREpTEV
Average holdup BI: -0
Normalized holdup BI: -0
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 4
Protein A protein: SNTA1
Protein A fragment: 84-171
Protein A site: PDZ
Protein A construct: his6-MBP-TEVsite-PDZ
Protein B protein: HPV35-E6
Protein B fragment: 140-149
Protein B site: PBM
Protein B sequence: SKPTRRETEV
Protein B construct: biotin-ttds-SKPTRRETEV
Average holdup BI: 0.47
Holdup BI standard deviation: 0.04
Normalized holdup BI: 0.54
Normalized holdup BI standard deviation: 0.04
Immobilized partner concentration in holdup experiment (10-6M): 18.55
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: 4.85
Experiment series 5
Protein A protein: SNTA1
Protein A fragment: 84-171
Protein A site: PDZ
Protein A construct: his6-MBP-TEVsite-PDZ
Protein B protein: HPV35-E6
Protein B fragment: 140-149
Protein B site: PBM
Protein B sequence: SKPTRRETEV
Protein B construct: biotin-ttds-SKPTRRETEV
Average holdup BI: 0.34
Normalized holdup BI: 0.44
Immobilized partner concentration in holdup experiment (10-6M): 18.55
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: 4.67
Experiment series 6
Protein A protein: SNTA1
Protein A fragment: 84-171
Protein A site: PDZ
Protein A construct: his6-MBP-TEVsite-PDZ
Protein B protein: HPV35-E6
Protein B fragment: 140-149
Protein B site: PBM
Protein B sequence: SKPTRRETEV
Protein B construct: biotin-ttds-SKPTRRETEV
Average holdup BI: 0.46
Normalized holdup BI: 0.52
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 #6
PUBMED ID of publication: 36115835
Number of measurements: 1
Measured pKd: 4.83