User:Lois A. Fridmann/Sandbox 1: Difference between revisions

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'''Green Flourscent Protein Mutant F99S//M153T/V163A'''<br>

'''with the ~~fluorescence~~ chromophore at the center of the barrel structure'''<br>

'''with the fluorescent chromophore at the center of the barrel structure'''<br>

'''accompanied by two water molecule and glutamate 222. The three'''<br>

'''mutant amino residues are found on the surface of the protein.'''

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==Background==

Considerable variations of the [[Green Fluorescent Protein]] (GFP) have been created in order to help determine the structure and function of the wtGFP. These mutations have been made in order to enhance the use of GFP as a reporter of gene regulation and expression, protein transport, and heredity. GFP has been mutated to provide an increased or alternative color for fluorescent, fluorescence for a greater time period and to make it less susceptible to photo bleaching. The specific GFP mutation ~~model created in RasMol and~~ featured in this page is referred to as F99S/M153T/V163A or cycle 3 GFP (c3-GFP).

Considerable variations of the [[Green Fluorescent Protein]] (GFP) have been created in order to help determine the structure and function of the wtGFP. These mutations have been made in order to enhance the use of GFP as a reporter of gene regulation and expression, protein transport and heredity. GFP has been mutated to provide an increased or alternative color for fluorescent, fluorescence for a greater time period and to make it less susceptible to photo bleaching. The specific GFP mutation featured in this page is referred to as F99S/M153T/V163A or cycle 3 GFP (c3-GFP).

The overall characteristic of the c3-GFP closely resemble the wtGFP, however due to the three mutations it is 42-fold more fluorescent in vivo compared to wtGFP. This phenomenon arises because the replacement of the hydrophobic amino acids with hydrophilic or less hydrophobic amino acids causes less aggregation and results in improved autocatalytic activation of the chromophore. The reduction in the hydrophobic nature on the surface of the c3-GFP also results in an increased ability of the mutant to mature in vivo effectively at 37 degrees Celsius. This is a feature the wtGFP lacks because the ''A. victoria'' jellyfish in the Pacific Northwest from which it was first extracted had never experienced such high temperatures~~, thus~~ the wtGFP has its limitations as a gene expression marker. The c3-GFP mutant is ~~special~~ because unlike most mutants, the structure of the chromophore and the fluorescence spectrum remains the same as the wtGFP. The enhanced c3-GFP provides for a better tool for researchers as compared with the wtGFP.

The overall characteristic of the c3-GFP closely resemble the wtGFP, however due to the three mutations it is 42-fold more fluorescent in vivo compared to wtGFP. This phenomenon arises because the replacement of the hydrophobic amino acids with hydrophilic or less hydrophobic amino acids causes less aggregation and results in improved autocatalytic activation of the chromophore. The reduction in the hydrophobic nature on the surface of the c3-GFP also results in an increased ability of the mutant to mature in vivo effectively at 37 degrees Celsius. This is a feature the wtGFP lacks because the ''A. victoria'' jellyfish in the Pacific Northwest, from which it was first extracted, had never experienced such high temperatures. Thus the wtGFP has its limitations as a gene expression marker. The c3-GFP mutant is unique because unlike most mutants, the structure of the chromophore and the fluorescence spectrum remains the same as the wtGFP. The enhanced c3-GFP provides for a better tool for researchers as compared with the wtGFP.

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==Notes and Literature References==

# Battistutta R, Negro A, Zanotti G. 2000. Crystal Structure and Refolding Properties of the Mutant F99S/M153T/V163A of the Green Fluorescent Protein. Proteins: Structure, Function, and Genetics 41:429 – 437.

@@ Line 4: / Line 4: @@
 '''Green Flourscent Protein Mutant F99S//M153T/V163A'''<br>
-'''with the fluorescence chromophore at the center of the barrel structure'''<br>
+'''with the fluorescent chromophore at the center of the barrel structure'''<br>
 '''accompanied by two water molecule and glutamate 222.  The three'''<br>
 '''mutant amino residues are found on the surface of the protein.'''
@@ Line 11: / Line 11: @@
 ==Background==
-Considerable variations of the [[Green Fluorescent Protein]] (GFP) have been created in order to help determine the structure and function of the wtGFP.  These mutations have been made in order to enhance the use of GFP as a reporter of gene regulation and expression, protein transport, and heredity.  GFP has been mutated to provide an increased or alternative color for fluorescent, fluorescence for a greater time period and to make it less susceptible to photo bleaching.  The specific GFP mutation model created in RasMol and featured in this page is referred to as F99S/M153T/V163A or cycle 3 GFP (c3-GFP).
+Considerable variations of the [[Green Fluorescent Protein]] (GFP) have been created in order to help determine the structure and function of the wtGFP.  These mutations have been made in order to enhance the use of GFP as a reporter of gene regulation and expression, protein transport and heredity.  GFP has been mutated to provide an increased or alternative color for fluorescent, fluorescence for a greater time period and to make it less susceptible to photo bleaching.  The specific GFP mutation featured in this page is referred to as F99S/M153T/V163A or cycle 3 GFP (c3-GFP).
 <applet load='1b9c' size='400' frame='true' align='right' caption='1b9c, resolution 2.40Å' />
-The overall characteristic of the c3-GFP closely resemble the wtGFP, however due to the three mutations it is 42-fold more fluorescent in vivo compared to wtGFP. This phenomenon arises because the replacement of the hydrophobic amino acids with hydrophilic or less hydrophobic amino acids causes less aggregation and results in improved autocatalytic activation of the chromophore. The reduction in the hydrophobic nature on the surface of the c3-GFP also results in an increased ability of the mutant to mature in vivo effectively at 37 degrees Celsius. This is a feature the wtGFP lacks because the ''A. victoria'' jellyfish in the Pacific Northwest from which it was first extracted had never experienced such high temperatures, thus the wtGFP has its limitations as a gene expression marker.  The c3-GFP mutant is special because unlike most mutants, the structure of the chromophore and the fluorescence spectrum remains the same as the wtGFP.  The enhanced c3-GFP provides for a better tool for researchers as compared with the wtGFP.
+The overall characteristic of the c3-GFP closely resemble the wtGFP, however due to the three mutations it is 42-fold more fluorescent in vivo compared to wtGFP. This phenomenon arises because the replacement of the hydrophobic amino acids with hydrophilic or less hydrophobic amino acids causes less aggregation and results in improved autocatalytic activation of the chromophore. The reduction in the hydrophobic nature on the surface of the c3-GFP also results in an increased ability of the mutant to mature in vivo effectively at 37 degrees Celsius. This is a feature the wtGFP lacks because the ''A. victoria'' jellyfish in the Pacific Northwest, from which it was first extracted, had never experienced such high temperatures.  Thus the wtGFP has its limitations as a gene expression marker.  The c3-GFP mutant is unique because unlike most mutants, the structure of the chromophore and the fluorescence spectrum remains the same as the wtGFP.  The enhanced c3-GFP provides for a better tool for researchers as compared with the wtGFP.
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@@ Line 22: / Line 22: @@
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 ==Notes and Literature References==
 # Battistutta R, Negro A, Zanotti G. 2000. Crystal Structure and Refolding Properties of the Mutant F99S/M153T/V163A of the Green Fluorescent Protein. Proteins: Structure, Function, and Genetics 41:429 – 437.