Interaction Details


Details of experiments between Q9Y4H2 and P63104
Note that the results of all experiments are listed, regardless of the modification states of the fragments.


Experiment series 1

Protein A

Protein: IRS2
Fragment: 1159-1168
Site: 14-3-3_pSer1164
Modification: (SEP)1164
Sequence: TPVSPSFAHN
Construct: TPVSP(pSer)FAHN (crude synthetic peptide)

Protein B

Protein: YWHAZ
Fragment: 1-245
Site: 14-3-3
Construct: avi-his6-MBP-TEVsite-14-3-3zeta
Average holdup BI: 0.01
Holdup BI standard deviation: 0.06
Immobilized partner concentration (10-6M): 132
P value: 0.16
Experiment method: HU Multiplex
Details of affinity fitting: hyperbolic binding equation
Date of measurement: 2021-10-13
Number of measurements: 5
The affinity was below the detection threshold of the assay.
DOI: 10.1101/2022.12.08.519103
Experimental Details

-log10(P Two-sided unpaired T-test (4 vs 5))

Experiment series 2

Protein A

Protein: IRS2
Fragment: 1171-1180
Site: 14-3-3_pSer1176
Modification: (SEP)1176
Sequence: RHNSASVENV
Construct: RHNSA(pSer)VENV (crude synthetic peptide)

Protein B

Protein: YWHAZ
Fragment: 1-245
Site: 14-3-3
Construct: avi-his6-MBP-TEVsite-14-3-3zeta
Average holdup BI: 0.15
Holdup BI standard deviation: 0.06
Immobilized partner concentration (10-6M): 132
P value: 2.33
Experiment method: HU Multiplex
Details of affinity fitting: hyperbolic binding equation
Date of measurement: 2021-10-13
Number of measurements: 5
The affinity was below the detection threshold of the assay.
DOI: 10.1101/2022.12.08.519103
Experimental Details

-log10(P Two-sided unpaired T-test (4 vs 5))

Experiment series 3

Protein A

Protein: IRS2
Fragment: 1278-1287
Site: 14-3-3_pSer1283
Modification: (SEP)1283
Sequence: PGDKSSWGRT
Construct: PGDKS(pSer)WGRT (crude synthetic peptide)

Protein B

Protein: YWHAZ
Fragment: 1-245
Site: 14-3-3
Construct: avi-his6-MBP-TEVsite-14-3-3zeta
Average holdup BI: 0
Holdup BI standard deviation: 0.09
Immobilized partner concentration (10-6M): 132
P value: 0.04
Experiment method: HU Multiplex
Details of affinity fitting: hyperbolic binding equation
Date of measurement: 2021-10-13
Number of measurements: 5
The affinity was below the detection threshold of the assay.
DOI: 10.1101/2022.12.08.519103
Experimental Details

-log10(P Two-sided unpaired T-test (4 vs 5))

Experiment series 4

Protein A

Protein: IRS2
Fragment: 341-350
Site: 14-3-3_pSer346
Modification: (SEP)346
Sequence: RSRTDSLAAT
Construct: RSRTD(pSer)LAAT (crude synthetic peptide)

Protein B

Protein: YWHAZ
Fragment: 1-245
Site: 14-3-3
Construct: avi-his6-MBP-TEVsite-14-3-3zeta
Average holdup BI: 0.02
Holdup BI standard deviation: 0.08
Immobilized partner concentration (10-6M): 132
P value: 0.23
Experiment method: HU Multiplex
Details of affinity fitting: hyperbolic binding equation
Date of measurement: 2021-10-13
Number of measurements: 5
The affinity was below the detection threshold of the assay.
DOI: 10.1101/2022.12.08.519103
Experimental Details

-log10(P Two-sided unpaired T-test (4 vs 5))

Experiment series 5

Protein A

Protein: IRS2
Fragment: 379-388
Site: 14-3-3_pSer384
Modification: (SEP)384
Sequence: GARPVSVAGS
Construct: GARPV(pSer)VAGS (crude synthetic peptide)

Protein B

Protein: YWHAZ
Fragment: 1-245
Site: 14-3-3
Construct: avi-his6-MBP-TEVsite-14-3-3zeta
Average holdup BI: 0.27
Holdup BI standard deviation: 0.03
Immobilized partner concentration (10-6M): 132
P value: 5.98
Experiment method: HU Multiplex
Details of affinity fitting: hyperbolic binding equation
Date of measurement: 2021-10-13
Number of measurements: 5
Measured pKd: 3.44
DOI: 10.1101/2022.12.08.519103
Experimental Details

-log10(P Two-sided unpaired T-test (4 vs 5))

Experiment series 6

Protein A

Protein: IRS2
Fragment: 427-436
Site: 14-3-3_pSer432
Modification: (SEP)432
Sequence: HSRSMSMPVA
Construct: HSRSM(pSer)MPVA (crude synthetic peptide)

Protein B

Protein: YWHAZ
Fragment: 1-245
Site: 14-3-3
Construct: avi-his6-MBP-TEVsite-14-3-3zeta
Average holdup BI: 0.88
Holdup BI standard deviation: 0.02
Immobilized partner concentration (10-6M): 132
P value: 6.72
Experiment method: HU Multiplex
Details of affinity fitting: hyperbolic binding equation
Date of measurement: 2021-10-13
Number of measurements: 5
Measured pKd: 4.72
DOI: 10.1101/2022.12.08.519103
Experimental Details

-log10(P Two-sided unpaired T-test (4 vs 5))

Experiment series 7

Protein A

Protein: IRS2
Fragment: 518-527
Site: 14-3-3_pSer523
Modification: (SEP)523
Sequence: SNTPESIAET
Construct: SNTPE(pSer)IAET (crude synthetic peptide)

Protein B

Protein: YWHAZ
Fragment: 1-245
Site: 14-3-3
Construct: avi-his6-MBP-TEVsite-14-3-3zeta
Average holdup BI: -0.06
Holdup BI standard deviation: 0.21
Immobilized partner concentration (10-6M): 132
P value: 0.31
Experiment method: HU Multiplex
Details of affinity fitting: hyperbolic binding equation
Date of measurement: 2021-10-13
Number of measurements: 5
The affinity was below the detection threshold of the assay.
DOI: 10.1101/2022.12.08.519103
Experimental Details

-log10(P Two-sided unpaired T-test (4 vs 5))

Experiment series 8

Protein A

Protein: IRS2
Fragment: 603-612
Site: 14-3-3_pSer608
Modification: (SEP)608
Sequence: ATFSGSAGRL
Construct: ATFSG(pSer)AGRL (crude synthetic peptide)

Protein B

Protein: YWHAZ
Fragment: 1-245
Site: 14-3-3
Construct: avi-his6-MBP-TEVsite-14-3-3zeta
Average holdup BI: 0.17
Holdup BI standard deviation: 0.03
Immobilized partner concentration (10-6M): 132
P value: 4.9
Experiment method: HU Multiplex
Details of affinity fitting: hyperbolic binding equation
Date of measurement: 2021-10-13
Number of measurements: 5
The affinity was below the detection threshold of the assay.
DOI: 10.1101/2022.12.08.519103
Experimental Details

-log10(P Two-sided unpaired T-test (4 vs 5))

Experiment series 9

Protein A

Protein: IRS2
Fragment: 718-727
Site: 14-3-3_pSer723
Modification: (SEP)723
Sequence: RTFPASGGGY
Construct: RTFPA(pSer)GGGY (crude synthetic peptide)

Protein B

Protein: YWHAZ
Fragment: 1-245
Site: 14-3-3
Construct: avi-his6-MBP-TEVsite-14-3-3zeta
Average holdup BI: 0.31
Holdup BI standard deviation: 0.02
Immobilized partner concentration (10-6M): 132
P value: 6.84
Experiment method: HU Multiplex
Details of affinity fitting: hyperbolic binding equation
Date of measurement: 2021-10-13
Number of measurements: 5
Measured pKd: 3.53
DOI: 10.1101/2022.12.08.519103
Experimental Details

-log10(P Two-sided unpaired T-test (4 vs 5))

Experiment series 10

Protein A

Protein: IRS2
Fragment: 725-734
Site: 14-3-3_pSer730
Modification: (SEP)730
Sequence: GGYKASSPAE
Construct: GGYKA(pSer)SPAE (crude synthetic peptide)

Protein B

Protein: YWHAZ
Fragment: 1-245
Site: 14-3-3
Construct: avi-his6-MBP-TEVsite-14-3-3zeta
Average holdup BI: -0.05
Holdup BI standard deviation: 0.12
Immobilized partner concentration (10-6M): 132
P value: 0.42
Experiment method: HU Multiplex
Details of affinity fitting: hyperbolic binding equation
Date of measurement: 2021-10-13
Number of measurements: 5
The affinity was below the detection threshold of the assay.
DOI: 10.1101/2022.12.08.519103
Experimental Details

-log10(P Two-sided unpaired T-test (4 vs 5))

Experiment series 11

Protein A

Protein: IRS2
Fragment: 574-583
Site: 14-3-3_pThr579
Modification: (TPO)579
Sequence: RTYSLTTPAR
Construct: RTYSL(pThr)TPAR (crude synthetic peptide)

Protein B

Protein: YWHAZ
Fragment: 1-245
Site: 14-3-3
Construct: avi-his6-MBP-TEVsite-14-3-3zeta
Measured dissociation constant (10-6M): <1
Immobilized partner concentration (10-6M): 132
Experiment method: HU Multiplex
Details of affinity fitting: hyperbolic binding equation
Date of measurement: 2021-10-13
Number of measurements: 1
Measured pKd: 6
DOI: 10.1101/2022.12.08.519103
Experimental Details

-log10(P Two-sided unpaired T-test (4 vs 5))

Experiment series 12

Protein A

Protein: IRS2
Fragment: 574-583
Site: 14-3-3_pThr579
Modification: (TPO)579
Sequence: RTYSLTTPAR
Construct: biotin-ttds-RTYSL(pThr)TPAR (HPLC-purified synthetic peptide)

Protein B

Protein: YWHAZ
Fragment: 1-245
Site: 14-3-3
Construct: avi-his6-MBP-TEVsite-14-3-3zeta
Measured dissociation constant (10-6M): 4.6
Standard deviation of dissociation constant (10-6M): 0.5
Experiment method: competitive Fluorescence Polarization
Details of affinity fitting: Competitive binding equation in ProFit
Date of measurement: 2022-02-10
Number of measurements: 3
Measured pKd: 5.33
DOI: 10.1101/2022.12.08.519103

Experiment series 13

Protein A

Protein: IRS2
Fragment: 574-583
Site: 14-3-3_pThr579
Modification: (TPO)579
Sequence: RTYSLTTPAR
Construct: biotin-ttds-RTYSL(pThr)TPAR (HPLC-purified synthetic peptide)

Protein B

Protein: YWHAZ
Fragment: 1-245
Site: 14-3-3
Construct: avi-his6-MBP-TEVsite-14-3-3zeta
Measured dissociation constant (10-6M): 6.3
Standard deviation of dissociation constant (10-6M): 0.4
Experiment method: competitive Fluorescence Polarization
Details of affinity fitting: Competitive binding equation in ProFit
Date of measurement: 2022-01-19
Number of measurements: 3
Measured pKd: 5.2
DOI: 10.1101/2022.12.08.519103