Negative control

Negative control prevents RNA polymerase from transcribing a gene through the use of regulatory proteins called "repressor proteins". In the active state, repressor proteins bind to DNA near the promoter region, and prevent RNA polymerase from transcribing that gene. In the inactive state, the repressor protein cannot bind to DNA, and the gene is transcribed.

Let's look at some examples.

Lactose Induction

E. coli can use a number of different sugars as energy sources, including lactose. To grow using lactose as a carbon source, E. coli must first transport it into the cell (using a lactose permease, the product of the lacY gene). Once inside, lactose is cleaved into glucose and galactose by an enzyme called B-galactosidase (the product of the lacZ gene).

The lacZ and lacY genes (along with another gene, lacA ) are organized into the Lac operon. In the absence of lactose, a repressor protein (the product of the lacI gene) sits near the promoter site, blocking RNA polymerase from transcribing the gene.

When lactose is present, lactose binds to the repressor protein, causing it to change into a conformation that can no longer bind to DNA, therefore allowing RNA polymerase to transcribe the genes. In this way, the genes that code enzymes that let the cell use lactose as an energy source are only made when lactose is present.

This is induction because the presence of the small molecule, lactose, causes the lactose utilization enzymes to be made.

What experiments can you do to show that induction is taking place ?

Let's look at another example.

Arginine Repression

Arginine is an essential amino acid for all cells. When there is plenty of arginine around, E. coli just brings it in through a transport protein and uses it from the external environment. However, when there is no arginine around, E. coli must make it from scratch, which requires a number of biosynthetic enzymes.

E. coli saves cellular resources by shutting off the genes for arginine biosynthesis when there is arginine around. E. coli only makes the enzymes for arginine biosynthesis when there is no arginine in the environment.

E. coli does this with a negative regulatory protein (a repressor protein) that binds to DNA near the promoter of the arginine genes. When there is no arginine present, the repressor is in its inactive conformation, and cannot bind to DNA. This means that the genes for arginine biosynthesis can be transcribed and translated.

However, when arginine IS present, the arginine binds to the repressor protein, causing the repressor protein to change to its active conformation. Now the repressor protein can bind to DNA near the promoter of the arginine genes, and stop transcription.

This is repression because the presence of the small molecule stops enzyme synthesis.

Now that you've read about it, watch the negative control animation!