Details of experiments between Q9H4L5 and P62258
Note that the results of all experiments are listed, regardless of the modification states of the fragments.
Experiment series 1
Protein A protein: OSBPL3
Protein A fragment: 299-308
Protein A site: 14-3-3_pSer304
Protein A modification: (SEP)304
Protein A sequence: VRLHSSNPNL
Protein A construct: VRLHS(pSer)NPNL (crude synthetic peptide)
Protein B protein: YWHAE
Protein B fragment: 1-255
Protein B site: 14-3-3
Protein B construct: avi-his6-MBP-TEVsite-14-3-3epsilon
Average holdup BI: 0.46
Holdup BI standard deviation: 0.01
Immobilized partner concentration in holdup experiment (10-6M): 116
P value (usually -log10(P Two-sided unpaired T-test), but double check the experimental details): 8.21
Experiment method: HU Multiplex
Details of affinity fitting: hyperbolic binding equation
Date of measurement: 2022.02.17.
Experimental details: -log10(P Two-sided unpaired T-test (4 vs 4))
Number of measurements: 4
Measured pKd: 3.87
Experiment series 2
Protein A protein: OSBPL3
Protein A fragment: 246-255
Protein A site: 14-3-3_pSer251
Protein A modification: (SEP)251
Protein A sequence: LHRTYSAPAI
Protein A construct: LHRTY(pSer)APAI (crude synthetic peptide)
Protein B protein: YWHAE
Protein B fragment: 1-255
Protein B site: 14-3-3
Protein B construct: avi-his6-MBP-TEVsite-14-3-3epsilon
Average holdup BI: 0.68
Holdup BI standard deviation: 0.03
Immobilized partner concentration in holdup experiment (10-6M): 143
P value (usually -log10(P Two-sided unpaired T-test), but double check the experimental details): 6.37
Experiment method: HU Multiplex
Details of affinity fitting: hyperbolic binding equation
Date of measurement: 2021.10.13.
Experimental details: -log10(P Two-sided unpaired T-test (4 vs 5))
Number of measurements: 5
Measured pKd: 4.17
Experiment series 3
Protein A protein: OSBPL3
Protein A fragment: 299-308
Protein A site: 14-3-3_pSer304
Protein A modification: (SEP)304
Protein A sequence: VRLHSSNPNL
Protein A construct: VRLHS(pSer)NPNL (crude synthetic peptide)
Protein B protein: YWHAE
Protein B fragment: 1-255
Protein B site: 14-3-3
Protein B construct: avi-his6-MBP-TEVsite-14-3-3epsilon
Average holdup BI: 0.56
Holdup BI standard deviation: 0.05
Immobilized partner concentration in holdup experiment (10-6M): 143
P value (usually -log10(P Two-sided unpaired T-test), but double check the experimental details): 5.76
Experiment method: HU Multiplex
Details of affinity fitting: hyperbolic binding equation
Date of measurement: 2021.10.13.
Experimental details: -log10(P Two-sided unpaired T-test (4 vs 5))
Number of measurements: 5
Measured pKd: 3.96
Experiment series 4
Protein A protein: OSBPL3
Protein A fragment: 29-38
Protein A site: 14-3-3_pSer34
Protein A modification: (SEP)34
Protein A sequence: GSRQDSWEVV
Protein A construct: GSRQD(pSer)WEVV (crude synthetic peptide)
Protein B protein: YWHAE
Protein B fragment: 1-255
Protein B site: 14-3-3
Protein B construct: avi-his6-MBP-TEVsite-14-3-3epsilon
Average holdup BI: 0.25
Holdup BI standard deviation: 0.26
Immobilized partner concentration in holdup experiment (10-6M): 116
P value (usually -log10(P Two-sided unpaired T-test), but double check the experimental details): 0.94
Experiment method: HU Multiplex
Details of affinity fitting: hyperbolic binding equation
Date of measurement: 2022.02.17.
Experimental details: -log10(P Two-sided unpaired T-test (4 vs 4))
Number of measurements: 4
The affinity was below the detection threshold of the assay.
Experiment series 5
Protein A protein: OSBPL3
Protein A fragment: 29-38
Protein A site: 14-3-3_pSer34
Protein A modification: (SEP)34
Protein A sequence: GSRQDSWEVV
Protein A construct: GSRQD(pSer)WEVV (crude synthetic peptide)
Protein B protein: YWHAE
Protein B fragment: 1-255
Protein B site: 14-3-3
Protein B construct: avi-his6-MBP-TEVsite-14-3-3epsilon
Average holdup BI: 0.02
Holdup BI standard deviation: 0.05
Immobilized partner concentration in holdup experiment (10-6M): 143
P value (usually -log10(P Two-sided unpaired T-test), but double check the experimental details): 0.43
Experiment method: HU Multiplex
Details of affinity fitting: hyperbolic binding equation
Date of measurement: 2021.10.13.
Experimental details: -log10(P Two-sided unpaired T-test (4 vs 5))
Number of measurements: 5
The affinity was below the detection threshold of the assay.
Experiment series 6
Protein A protein: OSBPL3
Protein A fragment: 432-441
Protein A site: 14-3-3_pSer437
Protein A modification: (SEP)437
Protein A sequence: LVHQLSNESR
Protein A construct: LVHQL(pSer)NESR (crude synthetic peptide)
Protein B protein: YWHAE
Protein B fragment: 1-255
Protein B site: 14-3-3
Protein B construct: avi-his6-MBP-TEVsite-14-3-3epsilon
Average holdup BI: -0.02
Holdup BI standard deviation: 0.06
Immobilized partner concentration in holdup experiment (10-6M): 143
P value (usually -log10(P Two-sided unpaired T-test), but double check the experimental details): 0.27
Experiment method: HU Multiplex
Details of affinity fitting: hyperbolic binding equation
Date of measurement: 2021.10.13.
Experimental details: -log10(P Two-sided unpaired T-test (4 vs 5))
Number of measurements: 5
The affinity was below the detection threshold of the assay.
Experiment series 7
Protein A protein: OSBPL3
Protein A fragment: 432-441
Protein A site: 14-3-3_pSer437
Protein A modification: (SEP)437
Protein A sequence: LVHQLSNESR
Protein A construct: LVHQL(pSer)NESR (crude synthetic peptide)
Protein B protein: YWHAE
Protein B fragment: 1-255
Protein B site: 14-3-3
Protein B construct: avi-his6-MBP-TEVsite-14-3-3epsilon
Average holdup BI: -0.02
Holdup BI standard deviation: 0.06
Immobilized partner concentration in holdup experiment (10-6M): 116
P value (usually -log10(P Two-sided unpaired T-test), but double check the experimental details): 0.23
Experiment method: HU Multiplex
Details of affinity fitting: hyperbolic binding equation
Date of measurement: 2022.02.17.
Experimental details: -log10(P Two-sided unpaired T-test (4 vs 4))
Number of measurements: 4
The affinity was below the detection threshold of the assay.
Experiment series 8
Protein A protein: OSBPL3
Protein A fragment: 315-324
Protein A site: 14-3-3_pSer320
Protein A modification: (SEP)320
Protein A sequence: EEKNYSDGSE
Protein A construct: EEKNY(pSer)DGSE (crude synthetic peptide)
Protein B protein: YWHAE
Protein B fragment: 1-255
Protein B site: 14-3-3
Protein B construct: avi-his6-MBP-TEVsite-14-3-3epsilon
Average holdup BI: -0.3
Holdup BI standard deviation: 1.43
Immobilized partner concentration in holdup experiment (10-6M): 116
P value (usually -log10(P Two-sided unpaired T-test), but double check the experimental details): 0.1
Experiment method: HU Multiplex
Details of affinity fitting: hyperbolic binding equation
Date of measurement: 2022.02.17.
Experimental details: -log10(P Two-sided unpaired T-test (4 vs 4))
Number of measurements: 4
The affinity was below the detection threshold of the assay.