Details of experiments between P50570 and Q14155
Note that the results of all experiments are listed, regardless of the modification states of the fragments.
Experiment series 1
Protein A protein: DNM2
Protein A fragment: 1-870
Protein A construct: endogenous protein (Jurkat)
Protein B protein: ARHGEF7
Protein B fragment: 178-251
Protein B site: SH3
Protein B construct: avi(biotin)-his6-MBP-TEVsite-SH3
Average holdup BI: 0.62
Immobilized partner concentration in holdup experiment (10-6M): 10
P value (usually -log10(P Two-sided unpaired T-test), but double check the experimental details): 8.07
Experiment method: nHU_Jurkat
Details of affinity fitting: hyperbolic binding equation
Date of measurement: 2022.06.29.
Experimental details: -log10(P Two-sided unpaired T-test (6 vs 3))
Number of measurements: 3
Measured pKd: 5.21
Experiment series 2
Protein A protein: DNM2
Protein A fragment: 313-327
Protein A site: PxxP_1
Protein A sequence: EYKNFRPDDPTRKTK
Protein A construct: Biotinylated 15-mer peptide (crude)
Protein B protein: ARHGEF7
Protein B fragment: 178-251
Protein B site: SH3
Protein B construct: avi(biotin)-his6-MBP-TEVsite-SH3
Average holdup BI: -0.01
Holdup BI standard deviation: 0
Immobilized partner concentration in holdup experiment (10-6M): 18
Experiment method: HU (DAPF)
Date of measurement: 2022.06.29.
Number of measurements: 2
The affinity was below the detection threshold of the assay.
Experiment series 3
Protein A protein: DNM2
Protein A fragment: 801-815
Protein A site: PxxP_2
Protein A sequence: AASFSAPPIPSRPGP
Protein A construct: Biotinylated 15-mer peptide (crude)
Protein B protein: ARHGEF7
Protein B fragment: 178-251
Protein B site: SH3
Protein B construct: avi(biotin)-his6-MBP-TEVsite-SH3
Average holdup BI: 0.25
Holdup BI standard deviation: 0.05
Immobilized partner concentration in holdup experiment (10-6M): 18
Experiment method: HU (DAPF)
Date of measurement: 2022.06.29.
Number of measurements: 2
Measured pKd: 4.3
Experiment series 4
Protein A protein: DNM2
Protein A fragment: 823-837
Protein A site: PxxP_3
Protein A sequence: DLFPAPPQIPSRPVR
Protein A construct: Biotinylated 15-mer peptide (crude)
Protein B protein: ARHGEF7
Protein B fragment: 178-251
Protein B site: SH3
Protein B construct: avi(biotin)-his6-MBP-TEVsite-SH3
Average holdup BI: 0.45
Holdup BI standard deviation: 0.05
Immobilized partner concentration in holdup experiment (10-6M): 18
Experiment method: HU (DAPF)
Date of measurement: 2022.06.29.
Number of measurements: 2
Measured pKd: 4.7
Experiment series 5
Protein A protein: DNM2
Protein A fragment: 826-840
Protein A site: PxxP_4
Protein A sequence: PAPPQIPSRPVRIPP
Protein A construct: Biotinylated 15-mer peptide (crude)
Protein B protein: ARHGEF7
Protein B fragment: 178-251
Protein B site: SH3
Protein B construct: avi(biotin)-his6-MBP-TEVsite-SH3
Average holdup BI: 0.41
Holdup BI standard deviation: 0.03
Immobilized partner concentration in holdup experiment (10-6M): 18
Experiment method: HU (DAPF)
Date of measurement: 2022.06.29.
Number of measurements: 2
Measured pKd: 4.64
Experiment series 6
Protein A protein: DNM2
Protein A fragment: 834-848
Protein A site: PxxP_5
Protein A sequence: RPVRIPPGIPPGVPS
Protein A construct: Biotinylated 15-mer peptide (crude)
Protein B protein: ARHGEF7
Protein B fragment: 178-251
Protein B site: SH3
Protein B construct: avi(biotin)-his6-MBP-TEVsite-SH3
Average holdup BI: 0.04
Holdup BI standard deviation: 0.01
Immobilized partner concentration in holdup experiment (10-6M): 18
Experiment method: HU (DAPF)
Date of measurement: 2022.06.29.
Number of measurements: 2
The affinity was below the detection threshold of the assay.
Experiment series 7
Protein A protein: DNM2
Protein A fragment: 838-852
Protein A site: PxxP_6
Protein A sequence: IPPGIPPGVPSRRPP
Protein A construct: Biotinylated 15-mer peptide (crude)
Protein B protein: ARHGEF7
Protein B fragment: 178-251
Protein B site: SH3
Protein B construct: avi(biotin)-his6-MBP-TEVsite-SH3
Average holdup BI: 0.15
Holdup BI standard deviation: 0.01
Immobilized partner concentration in holdup experiment (10-6M): 18
Experiment method: HU (DAPF)
Date of measurement: 2022.06.29.
Number of measurements: 2
Measured pKd: 3.99
Experiment series 8
Protein A protein: DNM2
Protein A fragment: 846-860
Protein A site: PxxP_7
Protein A sequence: VPSRRPPAAPSRPTI
Protein A construct: Biotinylated 15-mer peptide (crude)
Protein B protein: ARHGEF7
Protein B fragment: 178-251
Protein B site: SH3
Protein B construct: avi(biotin)-his6-MBP-TEVsite-SH3
Average holdup BI: 0.14
Holdup BI standard deviation: 0.04
Immobilized partner concentration in holdup experiment (10-6M): 18
Experiment method: HU (DAPF)
Date of measurement: 2022.06.29.
Number of measurements: 4
Measured pKd: 3.99
Experiment series 9
Protein A protein: DNM2
Protein A fragment: 826-840
Protein A site: PxxP_4
Protein A sequence: PAPPQIPSRPVRIPP
Protein A construct: Bio E E Ado Ado-PAPPQIPSRPVRIPP-CONH2
Protein B protein: ARHGEF7
Protein B fragment: 178-251
Protein B site: SH3
Protein B construct: his6-MBP-TEVsite-SH3
Average holdup BI: 0.38
Immobilized partner concentration in holdup experiment (10-6M): 18
Experiment method: HU (DAPF)
Details of affinity fitting: Quadratic binding equation
Date of measurement: 2023/01/17
Number of measurements: 1
Measured pKd: 4.57
Experiment series 10
Protein A protein: DNM2
Protein A fragment: 838-852
Protein A site: PxxP_6
Protein A sequence: IPPGIPPGVPSRRPP
Protein A construct: Bio E E Ado Ado-IPPGIPPGVPSRRPP-CONH2
Protein B protein: ARHGEF7
Protein B fragment: 178-251
Protein B site: SH3
Protein B construct: his6-MBP-TEVsite-SH3
Average holdup BI: 0.15
Immobilized partner concentration in holdup experiment (10-6M): 18
Experiment method: HU (DAPF)
Details of affinity fitting: Quadratic binding equation
Date of measurement: 2023/01/17
Number of measurements: 1
Measured pKd: 4