Details of experiments between the fragments P50570~823-860 and O00499~513-593
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: 823-860
Protein A construct: Biotinylated peptide
Protein B protein: BIN1
Protein B fragment: 513-593
Protein B site: SH3
Protein B modification: D537V
Protein B construct: avi(biotin)-his6-MBP-TEVsite-SH3
Average holdup BI: 0
Holdup BI standard deviation: 0.03
Immobilized partner concentration in holdup experiment (10-6M): 3
Experiment method: HU (DAPF)
Date of measurement: 2022.10.24.
Number of measurements: 18
The affinity was below the detection threshold of the assay.
Experiment series 2
Protein A protein: DNM2
Protein A fragment: 823-860
Protein A construct: Biotinylated peptide
Protein B protein: BIN1
Protein B fragment: 513-593
Protein B site: SH3
Protein B modification: F584S
Protein B construct: avi(biotin)-his6-MBP-TEVsite-SH3
Average holdup BI: -0.04
Holdup BI standard deviation: 0.05
Immobilized partner concentration in holdup experiment (10-6M): 3
Experiment method: HU (DAPF)
Date of measurement: 2022.10.24.
Number of measurements: 18
The affinity was below the detection threshold of the assay.
Experiment series 3
Protein A protein: DNM2
Protein A fragment: 823-860
Protein A construct: Biotinylated peptide
Protein B protein: BIN1
Protein B fragment: 513-593
Protein B site: SH3
Protein B modification: P551L
Protein B construct: avi(biotin)-his6-MBP-TEVsite-SH3
Average holdup BI: 0.18
Holdup BI standard deviation: 0.03
Immobilized partner concentration in holdup experiment (10-6M): 3
Experiment method: HU (DAPF)
Details of affinity fitting: Bait concentration was adjusted to fit the profile to WT
Date of measurement: 2022.10.24.
Number of measurements: 6
Measured pKd: 4.61
Experiment series 4
Protein A protein: DNM2
Protein A fragment: 823-860
Protein A construct: Biotinylated peptide
Protein B protein: BIN1
Protein B fragment: 513-593
Protein B site: SH3
Protein B modification: Q540H
Protein B construct: avi(biotin)-his6-MBP-TEVsite-SH3
Average holdup BI: 0.41
Holdup BI standard deviation: 0.02
Immobilized partner concentration in holdup experiment (10-6M): 3
Experiment method: HU (DAPF)
Date of measurement: 2022.10.24.
Number of measurements: 18
Measured pKd: 4.63
Experiment series 5
Protein A protein: DNM2
Protein A fragment: 823-860
Protein A construct: Biotinylated peptide
Protein B protein: BIN1
Protein B fragment: 513-593
Protein B site: SH3
Protein B modification: R581C
Protein B construct: avi(biotin)-his6-MBP-TEVsite-SH3
Average holdup BI: 0.19
Holdup BI standard deviation: 0.03
Immobilized partner concentration in holdup experiment (10-6M): 3
Experiment method: HU (DAPF)
Details of affinity fitting: Bait concentration was adjusted to fit the profile to WT
Date of measurement: 2022.10.24.
Number of measurements: 6
Measured pKd: 4.62
Experiment series 6
Protein A protein: DNM2
Protein A fragment: 823-860
Protein A construct: Biotinylated peptide
Protein B protein: BIN1
Protein B fragment: 513-593
Protein B site: SH3
Protein B modification: T532M
Protein B construct: avi(biotin)-his6-MBP-TEVsite-SH3
Average holdup BI: 0.34
Holdup BI standard deviation: 0.02
Immobilized partner concentration in holdup experiment (10-6M): 3
Experiment method: HU (DAPF)
Date of measurement: 2022.10.24.
Number of measurements: 18
Measured pKd: 4.48
Experiment series 7
Protein A protein: DNM2
Protein A fragment: 823-860
Protein A construct: Biotinylated peptide
Protein B protein: BIN1
Protein B fragment: 513-593
Protein B site: SH3
Protein B modification: V566M
Protein B construct: avi(biotin)-his6-MBP-TEVsite-SH3
Average holdup BI: 0.35
Holdup BI standard deviation: 0.02
Immobilized partner concentration in holdup experiment (10-6M): 3
Experiment method: HU (DAPF)
Date of measurement: 2022.10.24.
Number of measurements: 18
Measured pKd: 4.51
Experiment series 8
Protein A protein: DNM2
Protein A fragment: 823-860
Protein A construct: Biotinylated peptide
Protein B protein: BIN1
Protein B fragment: 513-593
Protein B site: SH3
Protein B modification: V583I
Protein B construct: avi(biotin)-his6-MBP-TEVsite-SH3
Average holdup BI: 0.27
Holdup BI standard deviation: 0.01
Immobilized partner concentration in holdup experiment (10-6M): 3
Experiment method: HU (DAPF)
Details of affinity fitting: Bait concentration was adjusted to fit the profile to WT
Date of measurement: 2022.10.24.
Number of measurements: 9
Measured pKd: 4.67
Experiment series 9
Protein A protein: DNM2
Protein A fragment: 823-860
Protein A construct: Biotinylated peptide
Protein B protein: BIN1
Protein B fragment: 513-593
Protein B site: SH3
Protein B construct: avi(biotin)-his6-MBP-TEVsite-SH3
Average holdup BI: 0.37
Holdup BI standard deviation: 0.01
Immobilized partner concentration in holdup experiment (10-6M): 3
Experiment method: HU (DAPF)
Date of measurement: 2022.10.24.
Number of measurements: 18
Measured pKd: 4.55
Experiment series 10
Protein A protein: DNM2
Protein A fragment: 823-860
Protein A construct: Biotinylated peptide
Protein B protein: BIN1
Protein B fragment: 513-593
Protein B site: SH3
Protein B modification: Y531S
Protein B construct: avi(biotin)-his6-MBP-TEVsite-SH3
Average holdup BI: 0.03
Holdup BI standard deviation: 0.02
Immobilized partner concentration in holdup experiment (10-6M): 3
Experiment method: HU (DAPF)
Date of measurement: 2022.10.24.
Number of measurements: 18
The affinity was below the detection threshold of the assay.