Details of experiments between the fragments O00499~513-593 and P50570~838-852
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: avi(biotin)-his6-MBP-TEVsite-SH3

Protein B protein: DNM2
Protein B fragment: 838-852
Protein B site: PxxP_6
Protein B sequence: IPPGIPPGVPSRRPP
Protein B construct: Biotinylated 15-mer peptide (crude)

Average holdup BI: 0.1
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
Measured pKd: 3.82


Experiment series 2

Protein A protein: BIN1
Protein A fragment: 513-593
Protein A site: SH3
Protein A modification: T532M
Protein A construct: avi(biotin)-his6-MBP-TEVsite-SH3

Protein B protein: DNM2
Protein B fragment: 838-852
Protein B site: PxxP_6
Protein B sequence: IPPGIPPGVPSRRPP
Protein B construct: Biotinylated 15-mer peptide (crude)

Average holdup BI: 0.1
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
Measured pKd: 3.79


Experiment series 3

Protein A protein: BIN1
Protein A fragment: 513-593
Protein A site: SH3
Protein A modification: D537V
Protein A construct: avi(biotin)-his6-MBP-TEVsite-SH3

Protein B protein: DNM2
Protein B fragment: 838-852
Protein B site: PxxP_6
Protein B sequence: IPPGIPPGVPSRRPP
Protein B construct: Biotinylated 15-mer peptide (crude)

Average holdup BI: -0.06
Holdup BI standard deviation: 0
Immobilized partner concentration in holdup experiment (10-6M): 18
Experiment method: HU (DAPF)
Date of measurement: 2022.10.24.
Number of measurements: 2
The affinity was below the detection threshold of the assay.


Experiment series 4

Protein A protein: BIN1
Protein A fragment: 513-593
Protein A site: SH3
Protein A modification: F584S
Protein A construct: avi(biotin)-his6-MBP-TEVsite-SH3

Protein B protein: DNM2
Protein B fragment: 838-852
Protein B site: PxxP_6
Protein B sequence: IPPGIPPGVPSRRPP
Protein B construct: Biotinylated 15-mer peptide (crude)

Average holdup BI: -0.08
Holdup BI standard deviation: 0.05
Immobilized partner concentration in holdup experiment (10-6M): 18
Experiment method: HU (DAPF)
Date of measurement: 2022.10.24.
Number of measurements: 2
The affinity was below the detection threshold of the assay.


Experiment series 5

Protein A protein: BIN1
Protein A fragment: 513-593
Protein A site: SH3
Protein A modification: P551L
Protein A construct: avi(biotin)-his6-MBP-TEVsite-SH3

Protein B protein: DNM2
Protein B fragment: 838-852
Protein B site: PxxP_6
Protein B sequence: IPPGIPPGVPSRRPP
Protein B construct: Biotinylated 15-mer peptide (crude)

Average holdup BI: 0.04
Holdup BI standard deviation: 0.03
Immobilized partner concentration in holdup experiment (10-6M): 6
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: 2
The affinity was below the detection threshold of the assay.


Experiment series 6

Protein A protein: BIN1
Protein A fragment: 513-593
Protein A site: SH3
Protein A modification: Q540H
Protein A construct: avi(biotin)-his6-MBP-TEVsite-SH3

Protein B protein: DNM2
Protein B fragment: 838-852
Protein B site: PxxP_6
Protein B sequence: IPPGIPPGVPSRRPP
Protein B construct: Biotinylated 15-mer peptide (crude)

Average holdup BI: 0.07
Holdup BI standard deviation: 0.03
Immobilized partner concentration in holdup experiment (10-6M): 18
Experiment method: HU (DAPF)
Date of measurement: 2022.10.24.
Number of measurements: 2
Measured pKd: 3.65


Experiment series 7

Protein A protein: BIN1
Protein A fragment: 513-593
Protein A site: SH3
Protein A modification: T532M
Protein A construct: avi(biotin)-his6-MBP-TEVsite-SH3

Protein B protein: DNM2
Protein B fragment: 838-852
Protein B site: PxxP_6
Protein B sequence: IPPGIPPGVPSRRPP
Protein B construct: Biotinylated 15-mer peptide (crude)

Average holdup BI: 0.05
Holdup BI standard deviation: 0
Immobilized partner concentration in holdup experiment (10-6M): 18
Experiment method: HU (DAPF)
Date of measurement: 2022.10.24.
Number of measurements: 2
Measured pKd: 3.48


Experiment series 8

Protein A protein: BIN1
Protein A fragment: 513-593
Protein A site: SH3
Protein A modification: V566M
Protein A construct: avi(biotin)-his6-MBP-TEVsite-SH3

Protein B protein: DNM2
Protein B fragment: 838-852
Protein B site: PxxP_6
Protein B sequence: IPPGIPPGVPSRRPP
Protein B construct: Biotinylated 15-mer peptide (crude)

Average holdup BI: 0.03
Holdup BI standard deviation: 0
Immobilized partner concentration in holdup experiment (10-6M): 18
Experiment method: HU (DAPF)
Date of measurement: 2022.10.24.
Number of measurements: 2
The affinity was below the detection threshold of the assay.


Experiment series 9

Protein A protein: BIN1
Protein A fragment: 513-593
Protein A site: SH3
Protein A modification: V583I
Protein A construct: avi(biotin)-his6-MBP-TEVsite-SH3

Protein B protein: DNM2
Protein B fragment: 838-852
Protein B site: PxxP_6
Protein B sequence: IPPGIPPGVPSRRPP
Protein B construct: Biotinylated 15-mer peptide (crude)

Average holdup BI: 0
Holdup BI standard deviation: 0.01
Immobilized partner concentration in holdup experiment (10-6M): 9
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: 2
The affinity was below the detection threshold of the assay.


Experiment series 10

Protein A protein: BIN1
Protein A fragment: 513-593
Protein A site: SH3
Protein A construct: avi(biotin)-his6-MBP-TEVsite-SH3

Protein B protein: DNM2
Protein B fragment: 838-852
Protein B site: PxxP_6
Protein B sequence: IPPGIPPGVPSRRPP
Protein B construct: Biotinylated 15-mer peptide (crude)

Average holdup BI: 0.08
Holdup BI standard deviation: 0.01
Immobilized partner concentration in holdup experiment (10-6M): 18
Experiment method: HU (DAPF)
Date of measurement: 2022.10.24.
Number of measurements: 2
Measured pKd: 3.69


Experiment series 11

Protein A protein: BIN1
Protein A fragment: 513-593
Protein A site: SH3
Protein A modification: Y531S
Protein A construct: avi(biotin)-his6-MBP-TEVsite-SH3

Protein B protein: DNM2
Protein B fragment: 838-852
Protein B site: PxxP_6
Protein B sequence: IPPGIPPGVPSRRPP
Protein B construct: Biotinylated 15-mer peptide (crude)

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: 2022.10.24.
Number of measurements: 2
The affinity was below the detection threshold of the assay.


Experiment series 12

Protein A protein: BIN1
Protein A fragment: 513-593
Protein A site: SH3
Protein A modification: R581C
Protein A construct: avi(biotin)-his6-MBP-TEVsite-SH3

Protein B protein: DNM2
Protein B fragment: 838-852
Protein B site: PxxP_6
Protein B sequence: IPPGIPPGVPSRRPP
Protein B construct: Biotinylated 15-mer peptide (crude)

Average holdup BI: 0.03
Holdup BI standard deviation: 0.01
Immobilized partner concentration in holdup experiment (10-6M): 6
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: 2
The affinity was below the detection threshold of the assay.


Experiment series 13

Protein A protein: BIN1
Protein A fragment: 513-593
Protein A site: SH3
Protein A construct: his6-MBP-TEVsite-SH3

Protein B protein: DNM2
Protein B fragment: 838-852
Protein B site: PxxP_6
Protein B sequence: IPPGIPPGVPSRRPP
Protein B construct: Bio E E Ado Ado-IPPGIPPGVPSRRPP-CONH2

Average holdup BI: 0.07
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: 3.6


Experiment series 14

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: DNM2
Protein B fragment: 838-852
Protein B site: PxxP_6
Protein B sequence: IPPGIPPGVPSRRPP
Protein B construct: Bio E E Ado Ado-IPPGIPPGVPSRRPP-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: 2023/01/16
Number of measurements: 1
Measured pKd: 3.75


Experiment series 15

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: DNM2
Protein B fragment: 838-852
Protein B site: PxxP_6
Protein B sequence: IPPGIPPGVPSRRPP
Protein B construct: Bio E E Ado Ado-IPPGIPPGVPSRRPP-CONH2

Average holdup BI: 0.08
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: 3.7


Experiment series 16

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: DNM2
Protein B fragment: 838-852
Protein B site: PxxP_6
Protein B sequence: IPPGIPPGVPSRRPP
Protein B construct: Bio E E Ado Ado-IPPGIPPGVPSRRPP-CONH2

Average holdup BI: 0.02
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 17

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: DNM2
Protein B fragment: 838-852
Protein B site: PxxP_6
Protein B sequence: IPPGIPPGVPSRRPP
Protein B construct: Bio E E Ado Ado-IPPGIPPGVPSRRPP-CONH2

Average holdup BI: 0.04
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
The affinity was below the detection threshold of the assay.


Experiment series 18

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: DNM2
Protein B fragment: 838-852
Protein B site: PxxP_6
Protein B sequence: IPPGIPPGVPSRRPP
Protein B construct: Bio E E Ado Ado-IPPGIPPGVPSRRPP-CONH2

Average holdup BI: 0.03
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
The affinity was below the detection threshold of the assay.


Experiment series 19

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: DNM2
Protein B fragment: 838-852
Protein B site: PxxP_6
Protein B sequence: IPPGIPPGVPSRRPP
Protein B construct: Bio E E Ado Ado-IPPGIPPGVPSRRPP-CONH2

Average holdup BI: -0.02
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 20

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: DNM2
Protein B fragment: 838-852
Protein B site: PxxP_6
Protein B sequence: IPPGIPPGVPSRRPP
Protein B construct: Bio E E Ado Ado-IPPGIPPGVPSRRPP-CONH2

Average holdup BI: 0.01
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
The affinity was below the detection threshold of the assay.


Experiment series 21

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: DNM2
Protein B fragment: 838-852
Protein B site: PxxP_6
Protein B sequence: IPPGIPPGVPSRRPP
Protein B construct: Bio E E Ado Ado-IPPGIPPGVPSRRPP-CONH2

Average holdup BI: 0.06
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: 3.83


Experiment series 22

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: DNM2
Protein B fragment: 838-852
Protein B site: PxxP_6
Protein B sequence: IPPGIPPGVPSRRPP
Protein B construct: Bio E E Ado Ado-IPPGIPPGVPSRRPP-CONH2

Average holdup BI: 0.08
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: 3.71