Details of experiments between the fragments O00499~513-593 and Q8IY67~420-434
Note that the results of all experiments are listed, regardless of the modification states of the fragments.
Experiment series 1
Protein A protein: BIN1
Protein A fragment: 513-593
Protein A site: SH3
Protein A construct: his6-MBP-TEVsite-SH3
Protein B protein: RAVER1
Protein B fragment: 420-434
Protein B site: PxxP_1
Protein B sequence: AGGGLPPELPPRRGK
Protein B construct: Bio Ado Ado-AGGGLPPELPPRRGK-CONH2
Average holdup BI: 0.56
Immobilized partner concentration in holdup experiment (10-6M): 18
Experiment method: HU (DAPF)
Details of affinity fitting: Quadratic binding equation
Date of measurement: 2023/01/16
Number of measurements: 1
Measured pKd: 4.91
Experiment series 2
Protein A protein: BIN1
Protein A fragment: 513-593
Protein A site: SH3
Protein A modification: T532M
Protein A construct: his6-MBP-TEVsite-SH3
Protein B protein: RAVER1
Protein B fragment: 420-434
Protein B site: PxxP_1
Protein B sequence: AGGGLPPELPPRRGK
Protein B construct: Bio Ado Ado-AGGGLPPELPPRRGK-CONH2
Average holdup BI: 0.57
Immobilized partner concentration in holdup experiment (10-6M): 18
Experiment method: HU (DAPF)
Details of affinity fitting: Quadratic binding equation
Date of measurement: 2023/01/16
Number of measurements: 1
Measured pKd: 4.92
Experiment series 3
Protein A protein: BIN1
Protein A fragment: 513-593
Protein A site: SH3
Protein A modification: Q540H
Protein A construct: his6-MBP-TEVsite-SH3
Protein B protein: RAVER1
Protein B fragment: 420-434
Protein B site: PxxP_1
Protein B sequence: AGGGLPPELPPRRGK
Protein B construct: Bio Ado Ado-AGGGLPPELPPRRGK-CONH2
Average holdup BI: 0.6
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: 4.97
Experiment series 4
Protein A protein: BIN1
Protein A fragment: 513-593
Protein A site: SH3
Protein A modification: D537V
Protein A construct: his6-MBP-TEVsite-SH3
Protein B protein: RAVER1
Protein B fragment: 420-434
Protein B site: PxxP_1
Protein B sequence: AGGGLPPELPPRRGK
Protein B construct: Bio Ado Ado-AGGGLPPELPPRRGK-CONH2
Average holdup BI: 0.09
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: 3.77
Experiment series 5
Protein A protein: BIN1
Protein A fragment: 513-593
Protein A site: SH3
Protein A modification: V583I
Protein A construct: his6-MBP-TEVsite-SH3
Protein B protein: RAVER1
Protein B fragment: 420-434
Protein B site: PxxP_1
Protein B sequence: AGGGLPPELPPRRGK
Protein B construct: Bio Ado Ado-AGGGLPPELPPRRGK-CONH2
Average holdup BI: 0.4
Immobilized partner concentration in holdup experiment (10-6M): 13
Experiment method: HU (DAPF)
Details of affinity fitting: Quadratic binding equation
Date of measurement: 2023/01/17
Number of measurements: 1
Measured pKd: 4.77
Experiment series 6
Protein A protein: BIN1
Protein A fragment: 513-593
Protein A site: SH3
Protein A modification: P551L
Protein A construct: his6-MBP-TEVsite-SH3
Protein B protein: RAVER1
Protein B fragment: 420-434
Protein B site: PxxP_1
Protein B sequence: AGGGLPPELPPRRGK
Protein B construct: Bio Ado Ado-AGGGLPPELPPRRGK-CONH2
Average holdup BI: 0.33
Immobilized partner concentration in holdup experiment (10-6M): 8
Experiment method: HU (DAPF)
Details of affinity fitting: Quadratic binding equation
Date of measurement: 2023/01/17
Number of measurements: 1
Measured pKd: 4.84
Experiment series 7
Protein A protein: BIN1
Protein A fragment: 513-593
Protein A site: SH3
Protein A modification: F584S
Protein A construct: his6-MBP-TEVsite-SH3
Protein B protein: RAVER1
Protein B fragment: 420-434
Protein B site: PxxP_1
Protein B sequence: AGGGLPPELPPRRGK
Protein B construct: Bio Ado Ado-AGGGLPPELPPRRGK-CONH2
Average holdup BI: -0.04
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
The affinity was below the detection threshold of the assay.
Experiment series 8
Protein A protein: BIN1
Protein A fragment: 513-593
Protein A site: SH3
Protein A modification: Y531S
Protein A construct: his6-MBP-TEVsite-SH3
Protein B protein: RAVER1
Protein B fragment: 420-434
Protein B site: PxxP_1
Protein B sequence: AGGGLPPELPPRRGK
Protein B construct: Bio Ado Ado-AGGGLPPELPPRRGK-CONH2
Average holdup BI: 0.24
Immobilized partner concentration in holdup experiment (10-6M): 18
Experiment method: HU (DAPF)
Details of affinity fitting: Quadratic binding equation
Date of measurement: 2023/01/18
Number of measurements: 1
Measured pKd: 4.26
Experiment series 9
Protein A protein: BIN1
Protein A fragment: 513-593
Protein A site: SH3
Protein A modification: R581C
Protein A construct: his6-MBP-TEVsite-SH3
Protein B protein: RAVER1
Protein B fragment: 420-434
Protein B site: PxxP_1
Protein B sequence: AGGGLPPELPPRRGK
Protein B construct: Bio Ado Ado-AGGGLPPELPPRRGK-CONH2
Average holdup BI: 0.37
Immobilized partner concentration in holdup experiment (10-6M): 10
Experiment method: HU (DAPF)
Details of affinity fitting: Quadratic binding equation
Date of measurement: 2023/01/18
Number of measurements: 1
Measured pKd: 4.83
Experiment series 10
Protein A protein: BIN1
Protein A fragment: 513-593
Protein A site: SH3
Protein A modification: V566M
Protein A construct: his6-MBP-TEVsite-SH3
Protein B protein: RAVER1
Protein B fragment: 420-434
Protein B site: PxxP_1
Protein B sequence: AGGGLPPELPPRRGK
Protein B construct: Bio Ado Ado-AGGGLPPELPPRRGK-CONH2
Average holdup BI: 0.59
Immobilized partner concentration in holdup experiment (10-6M): 18
Experiment method: HU (DAPF)
Details of affinity fitting: Quadratic binding equation
Date of measurement: 2023/01/18
Number of measurements: 1
Measured pKd: 4.96