(Transcribed from Dr. Cadilla’s lecture, 22 Mar 2000 by Brian Buschman)
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RNA comes in three main forms:
1) rRNA (ribosomal) where the most abundant and actually carry out translation of mRNA.
2) tRNA (transcriptional) which function to bind amino acids and bring them to the rRNA as they are coded for.
3) mRNA (messenger) are the RNA strands that will be transcribed into proteins.
Structurally RNA is very similar to DNA in that it is made from a chain of nucleotides and is synthesized 5’ to 3’ and read 3’ to 5’. It differs in that the sugar is ribose rather then deoxyribose and it is single stranded. It also uses U as the base in the place of T.
Bacteria only have one RNA polymerase which has a core enzyme made of subunits a2, b and b’ that requires the s subunit to initiate transcription. The RNA polymerase finds and binds to the promoter upstream. At first it weakly binds to the promoter until the s subunit comes along and “locks” the polymerase to the promoter.
The RNA polymerase knows where to start because it recognizes the promoter. There are usually two main promoter sequences the first called the –35 sequence (because it is located 35 bases before transcription starts) and the main promoter called the Pribnow Box (-10 box or TATAAT box).
When the polymerase is bound to the s subunit locks it to the promoter. Then helicase comes in and unwinds 18 bases before transcription starts. Then the s subunits disassociates and RNA polymerase does it’s thing. The elongation rate of the process varies with the DNA sequence. The more similar the general sequence of the gene is to the promoter the faster transcription will go.
In a manner similar to DNA replication RNA synthesis uses nucleotide triphosphates (ATP, TTP, GTP, CTP). One major difference is that RNA polymerase has no proofreading ability. That is fine because error rates are low and when an error does occur in transcription it causes few problems.
Transcription will continue until it’s end is signaled one of two was. First the r-factor can cause termination and second it can reach a palindrome sequence. A palindrome is a symmetrical sequence that will form a double strand with itself therefore forming a two dimensional DNA strand.
Many antibiotics function by effecting transcription such at:
Post translation modifications differ between eukaryotic and prokaryotic organisms. In the eukaryotic there are more cases of methylation of bases, glycolyslation of phosphates and the addition of the 5’ cap and 3’ poly-A tail. This all happens to eukaryotic mRNA.
Cells transcribe rRNA units as one strand and then cleave then into the two rRNAs cells need. This insures that equal numbers of both units exist in the cell.
In eukaryotes there are three different RNA polymerases:
1) RNA pol I makes rRNA.
2) RNA pol II makes mRNA.
3) RNA pol III makes tRNA.
Eukaryotes have a different set of promoters the main one being the Hogness box (TATA box) which is at about –25 and similar to the Pribnow box. There is also the CAAT box at about –75. Both serve as promoter sequences and the site for binding of transcription factors. For transcription to begin in the eukaryotic cell a number of transcription factors must bind to the DNA, pol II and to each other. They include TFIIA, TFIIB, TFIID, TFIIF, TFIIH and TFIII.
Enhancers are regions located on the DNA either upstream or downstream from the promoter that encourage the binding of transcription factors and increase the rate of transcription.
Inhibitors are similar to enhancers but they slow down the rate of transcription.
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