Based on the data shown where is the DNA binding domain?  Explain which constructs helped you reach this conclusion?       Which part of the protein is the Activation domain? Explain which constructs helped you reach your conclusion?

Human Anatomy & Physiology (11th Edition)
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  1. Based on the data shown where is the DNA binding domain?  Explain which constructs helped you reach this conclusion?

 

 

 

  1. Which part of the protein is the Activation domain? Explain which constructs helped you reach your conclusion? 
The schematic (a)below is that of a reporter-gene construct - containing a lacz reporter gene (encoding
B-galactosidase) and a TATA box ligated to UASGAL, a regulatory element that contains several binding
sites for transcription factor Gal4.
(a) Reporter-gene construct
(b) Expression Vector construct: Various plasmids containing coding sequences of wild-type or mutant
(portions of coding sequence deleted) Gal4 were made. (Plasmid 1 in diagram below)
Gene encoding
protein X
Both the reported gene construct and plasmids with DNA encoding wild-type or mutant (deleted) Gal4
were simultaneously introduced into mutant (gal4) yeast cells. (See schematic below)
Plasmid
1
2 activities were measured: In vitro binding (+ or -) to the reporter using EMSA's and the ability of
truncated proteins to activate transcription via production of reporter gene (B-gal activity) levels.
X-binding
site
Nucleus
Di
Reporter
UAS GAL
gene
Plasmid
Protein X
TATA
box
Reporter-gene
transcripts
(b) Wild-type and mutant Gal4 proteins
Binding
to UAS GAL
Wild type
Deletion
constructs
1
2
3
4
lacz gene
5
50
1 74
1 74
881
692
881
755
738881
B-Galactosidase
activity
+++
+++
Transcribed Image Text:The schematic (a)below is that of a reporter-gene construct - containing a lacz reporter gene (encoding B-galactosidase) and a TATA box ligated to UASGAL, a regulatory element that contains several binding sites for transcription factor Gal4. (a) Reporter-gene construct (b) Expression Vector construct: Various plasmids containing coding sequences of wild-type or mutant (portions of coding sequence deleted) Gal4 were made. (Plasmid 1 in diagram below) Gene encoding protein X Both the reported gene construct and plasmids with DNA encoding wild-type or mutant (deleted) Gal4 were simultaneously introduced into mutant (gal4) yeast cells. (See schematic below) Plasmid 1 2 activities were measured: In vitro binding (+ or -) to the reporter using EMSA's and the ability of truncated proteins to activate transcription via production of reporter gene (B-gal activity) levels. X-binding site Nucleus Di Reporter UAS GAL gene Plasmid Protein X TATA box Reporter-gene transcripts (b) Wild-type and mutant Gal4 proteins Binding to UAS GAL Wild type Deletion constructs 1 2 3 4 lacz gene 5 50 1 74 1 74 881 692 881 755 738881 B-Galactosidase activity +++ +++
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