Talk:20.109(S13):Assess protein function (Day8)

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|Comments (e.g., very low fluorescence)
|Comments (e.g., very low fluorescence)
|-
|-
-
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+
| Plt
-
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| M124S
-
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| D80R
-
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| WT: 0.0976; 1.4751
-
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| M124S: 0.4115; 3.9341
-
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| D80R: 1.2991; 2.8465
-
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| small difference between 0% and 100% for our mutant, affinity decrease and cooperativity increase
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|-
|Orange
|Orange
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|Very low fluorescence; oscillating trend...
|Very low fluorescence; oscillating trend...
|-
|-
-
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+
|Purple
-
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|M124S
-
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+
|M144R
-
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|KD = 0.0672, n = 1.1286
-
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+
|KD = 0.2682, n= 3.3655
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+
|KD = 0.3583, n= 4.1618
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|Higher cooperativity, lower affinity in target mutant
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|-
-
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|Blue
-
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+
|M124S
-
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+
|M71R
-
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|KD = 0.1175, n = 1.4880
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|KD = 0.3377, n = 3.3863
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|KD = 0.2246, n = 0.6741
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|Negative cooperativity, lower affinity in target mutant
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|-
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|Yellow
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|T79P
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|K77E
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|Kd = 0.1531, n = 1.4785
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|Kd = 0.8493, n = 1.5848
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|Kd = 0.0382, n = 1.1695
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|Design mutant: higher affinity, lower positive cooperativity
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|-
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|Green
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|T79P
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|M71R
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|Kd = 0.117, n = 1.50
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|Kd = 0.889, n = 4.05
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|Kd = 0.229, n = 0.684
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|T79P showed initial, slight increase in fluorescence, then sharp decrease. M71R showed sharply decreased cooperativity and increased Kd compared to WT.
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|-
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+
|Red
-
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+
|D24H
 +
|G25Q
 +
|Kd = 0.1232, n = 1.3583
 +
|Kd = 0.7541, n = 3.9963
 +
|Kd = 0.0415, n = -0.4091
 +
|Fluorescence rises to a peak then falls off at end
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Line 112: Line 117:
==Analysis, W/F==
==Analysis, W/F==
-
==Analysis, T/R==
+
==Analysis, W/F==
Please be mindful of significant figures when you fill in this table. Do not state far more precise numbers than you can believe.
Please be mindful of significant figures when you fill in this table. Do not state far more precise numbers than you can believe.
Line 125: Line 130:
|Comments (e.g., very low fluorescence)
|Comments (e.g., very low fluorescence)
|-
|-
-
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+
| Blue
-
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+
| D24H
-
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+
| D22E
-
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+
| Kd= 0.0781, n= 1.2420
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+
| Kd= 0.5804, n= 2.3194
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| Kd= 0.6807, n= 7.0509
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| Affinity decreased while the hill coefficient increased significantly. No extreme outliers.
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|-
-
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+
|Platinum
-
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| E67K
-
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| R126P
-
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| Kd = 0.0970 n =1.0639
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| Kd =  0.3729 n = 0.3830
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| Kd =  0.3488 n= 0.9085
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Low fluorescent values
|-
|-
-
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+
|Pink
-
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+
|D24H
-
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+
|G25Q
-
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+
|Kd = 0.1232, n = 1.3583
-
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+
|Kd = 0.7541, n = 3.9963
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|Kd = 0.0415, n = -0.4091
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|Low fluorescence relative to wt,  Affinity increased significantly or IPC was unable to refold properly.
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|-
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+
|Purple
-
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|M12S
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|M51T
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|Kd=0.1276, n=1.4726
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|Kd=0.3430, n=3.009
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|Kd=0.1191, n=1.2544
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|-
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|Yellow
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|M124S
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|M51T
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|Kd = 0.0739; n = 1.2038
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|Kd = 0.3859; n = 3.0814
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|Kd = 0.1403; n = 1.6126
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|Higher fluorescence compared to WT. Similar affinity as WT, higher affinity than reference.
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|-
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|Red
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|D24H
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|D22E
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|Kd = 0.0919, n = 1.858
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|Kd = 0.7099, n = 6.5600
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|Kd = 0.6134, n = 2.3619
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|Orange
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|D24H
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|D22E
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|Kd = 0.0916, n = 1.2896
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|Kd = 0.7357, n = 4.3991
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|Kd = 0.6766, n = 7.2874
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|Decreased affinity, increased cooperativity
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|Green
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|T79P
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|K77E
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|Kd = 6.75e-8M, n = 1.32
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|Kd = 9.30e-7M, n = 5.34
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|Kd = 1.84e-8M, n = 1.08
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|}

Current revision

Contents

Raw Data

Raw calcium titration data is posted below.


Day 7 data file, T/R Day 7 data file, W/F
Media:S13-M2D7-TR-Red.txt Media:S13-M2D7-WF-Red.txt
Media:S13-M2D7-TR-Org.txt Media:S13-M2D7-WF-Org.txt
Media:S13-M2D7-TR-Ylw.txt Media:S13-M2D7-WF-Ylw.txt
Media:S13-M2D7-TR-Grn.txt Media:S13-M2D7-WF-Grn.txt
Media:S13-M2D7-TR-Blu.txt Media:S13-M2D7-WF-Blu.txt
N/A Media:S13-M2D7-WF-Pnk.txt
Media:S13-M2D7-TR-Prp.txt Media:S13-M2D7-WF-Prp.txt
Media:S13-M2D7-TR-Plt.txt Media:S13-M2D7-WF-Plt.txt


Analysis, T/R

Please be mindful of significant figures when you fill in this table. Do not state far more precise numbers than you can believe.

Group Colour D24H, E67K, T79P, or M124S? X#Z Mutant Most believable KD; n for WT Most believable KD; n for reference Most believable KD; n for X#Z mutant Comments (e.g., very low fluorescence)
Plt M124S D80R WT: 0.0976; 1.4751 M124S: 0.4115; 3.9341 D80R: 1.2991; 2.8465 small difference between 0% and 100% for our mutant, affinity decrease and cooperativity increase
Orange D24H G25Q 0.1462, n = 0.8310 0.6759, n = 2.5734 0.7118; n = -0.2595 Very low fluorescence; oscillating trend...
Purple M124S M144R KD = 0.0672, n = 1.1286 KD = 0.2682, n= 3.3655 KD = 0.3583, n= 4.1618 Higher cooperativity, lower affinity in target mutant
Blue M124S M71R KD = 0.1175, n = 1.4880 KD = 0.3377, n = 3.3863 KD = 0.2246, n = 0.6741 Negative cooperativity, lower affinity in target mutant
Yellow T79P K77E Kd = 0.1531, n = 1.4785 Kd = 0.8493, n = 1.5848 Kd = 0.0382, n = 1.1695 Design mutant: higher affinity, lower positive cooperativity
Green T79P M71R Kd = 0.117, n = 1.50 Kd = 0.889, n = 4.05 Kd = 0.229, n = 0.684 T79P showed initial, slight increase in fluorescence, then sharp decrease. M71R showed sharply decreased cooperativity and increased Kd compared to WT.
Red D24H G25Q Kd = 0.1232, n = 1.3583 Kd = 0.7541, n = 3.9963 Kd = 0.0415, n = -0.4091 Fluorescence rises to a peak then falls off at end

Analysis, W/F

Analysis, W/F

Please be mindful of significant figures when you fill in this table. Do not state far more precise numbers than you can believe.

Group Colour D24H, E67K, T79P, or M124S? X#Z Mutant Most believable KD; n for WT Most believable KD; n for reference Most believable KD; n for X#Z mutant Comments (e.g., very low fluorescence)
Blue D24H D22E Kd= 0.0781, n= 1.2420 Kd= 0.5804, n= 2.3194 Kd= 0.6807, n= 7.0509 Affinity decreased while the hill coefficient increased significantly. No extreme outliers.
Platinum E67K R126P Kd = 0.0970 n =1.0639 Kd = 0.3729 n = 0.3830 Kd = 0.3488 n= 0.9085

Low fluorescent values

Pink D24H G25Q Kd = 0.1232, n = 1.3583 Kd = 0.7541, n = 3.9963 Kd = 0.0415, n = -0.4091 Low fluorescence relative to wt, Affinity increased significantly or IPC was unable to refold properly.
Purple M12S M51T Kd=0.1276, n=1.4726 Kd=0.3430, n=3.009 Kd=0.1191, n=1.2544
Yellow M124S M51T Kd = 0.0739; n = 1.2038 Kd = 0.3859; n = 3.0814 Kd = 0.1403; n = 1.6126 Higher fluorescence compared to WT. Similar affinity as WT, higher affinity than reference.
Red D24H D22E Kd = 0.0919, n = 1.858 Kd = 0.7099, n = 6.5600 Kd = 0.6134, n = 2.3619
Orange D24H D22E Kd = 0.0916, n = 1.2896 Kd = 0.7357, n = 4.3991 Kd = 0.6766, n = 7.2874 Decreased affinity, increased cooperativity
Green T79P K77E Kd = 6.75e-8M, n = 1.32 Kd = 9.30e-7M, n = 5.34 Kd = 1.84e-8M, n = 1.08
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