1、Major Contents of Lecture 5,Compare replication and transcription Operon structural gene, promoter RNA polymerase of E. Coli Transcription in prokaryotes,RNA biosynthesis takes place in transcription as well as RNA replication.,For some RNA viruses,The synthesis of RNA molecules using DNA strands as
2、 the templates is called transcription, by which the genetic information can be transferred from DNA to RNA.,Transcription,Question,Which molecules are included in the transcription system? NTP dNTP Primer Template RNA polymerase Ions,Compare Replication and Transcription,regulatory region,Operon Th
3、e unit of coordinated and regulated gene transcription. Its consist of structural genes and the regulatory region.,Sense (+),Structural gene Any gene that carries the genetic information for the structure of a protein.,translation,N - Ala Val His Val - C Protein,template strand,coding strand,templat
4、e strand,coding strand,In one gene, only the template strand but not the coding strand is used as the temple for RNA synthesis. Both strands of DNA can be used as templates.,Transcriptions of DNA are asymmetric.,Promoter A DNA sequence located 5 to a gene that indicates the initiation site for trans
5、cription.,Prokaryotic promoters may be determined from RNA-Pol Protection Assay for the reason that they can be recognized and bound by RNA-Pol.,regulatory region,Two Consensus Sequences in Prokaryotic Promoter,The promoter sequence may influence transcription activity of genes.,Frequency in 45 samp
6、les 38 36 29 40 25 3037 37 28 41 29 44,Fill in the Brackets,The ( ) strand of gene is used to direct RNA synthesis. If only one strand of DNA sequence is given, this strand usually is the ( ) strand. The ( ) strand is also called sense strand and plus strand. Pribnow box of prokaryotic promoters res
7、ides -10 region with a sequence of ( ). And TTGACA box resides ( ) region.,The subunit only exists in the holoenzyme but not in the core enzyme.,The core enzyme2 The holoenzyme2 ,RNA polymerase of E. coli,RNA Pol of E. coli,Functions of RNA Pol. Subunits,The 3-hydroxyl group nucleophilically attacks
8、 the phosphoryl group of the incoming nucleoside triphosphate. This transition state is structurally similar to that in DNA polymerase,A Model of the Active Site - the Transition State for Phosphodiester Bond Formation,Question,Is the fidelity of RNA synthesis as important as that of DNA replication
9、? Why?,Proofreading of RNA Pol Which Can Be Enhanced by Accessory Proteins.,The error rate 10-4 to 10-5 This fidelity is tolerable because of short lengths of RNA. In addition, mutations of RNA Not transmit to the progeny.No effect in some cases for being non-coding or non-sense.Not harmful in most
10、cases for many copies and fast degradation.,RifampicinA drug for tuberculosis treatment by inhibiting the initiation of RNA synthesis. It binds to the channel of RNA-pol for DNA-RNA hybrid pass (after two or three nucleotides connected).,The structure of the E. coli subunit 70 reveals an helix plays
11、 an important role in binding to Pribnow box.,70 interacts with -10 and -35 elements specifically.,Transcription in Prokaryotes,Initiation needs holoenzyme of RNA Polymerase and elongation needs the core enzyme. Interactions between promoter and subunit determine the start site of transcription. Tra
12、nslation to protein may occur before the mRNA synthesis finish. Termination may be factor dependent or factor independent.,Transcription of Prokaryotes,Initiation Stage Of Prokaryotic Transcription,The RNA-pol holoenzyme binds the TTGACA region and the TATAAT region, and then opens the DNA duplex.,T
13、ranscription initiation complex,RNA-pol (2) - DNA - pppGpN- OH 3,The first nucleotide on RNA transcript is always pppG or pppA.,No primer is required for RNA synthesis.,Elongation Stage of Prokaryotic Transcription,Release of the subunit The conformational change of core enzyme and the slide on the
14、DNA template forward. Free NTPs are added sequentially to the 3 -OH of the nascent RNA strand.,Core RNA-pol, DNA segment of 40nt and the nascent RNA form a complex called the transcription complex or the transcription bubble.,12bp,Multiple transcriptions can take place simultaneously.,Translation ma
15、y start in the elongation stage of prokaryotic transcription.,The protein binds to the C-rich region of nascent RNA, then play its roles of ATPase and helicase to catch the RNA pol. and release RNA transcript., protein,-Dependent Termination,Discovered by J. Roberts in 1969,-Independent Termination,
16、The termination signal of the RNA GC-rich and able to form stem-loop structures followed by a series of U,Terms and Concepts,transcriptionoperonstructrual genepromoterPribnow box subunit protein,Questions,Which similarities and difference are there between DNA replication and transcription? Please describe the structure and functions of Prokaryotic RNA polymerase. How is the transcription of E. coli terminated?,
