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<Structure load='3SPA' size='250' frame='true' align='right' caption='Human mitochondrial RNA polymerase' scene='Insert optional scene name here' />
<Structure load='3SPA' size='370' frame='true' align='right' caption='Human mitochondrial RNA polymerase (PDB: 3SPA) scene='Insert optional scene name here' />


One of the [[CBI Molecules]] being studied in the  [http://www.umass.edu/cbi/ University of Massachusetts Amherst Chemistry-Biology Interface Program] at UMass Amherst and on display at the [http://www.molecularplayground.org/ Molecular Playground].
One of the [[CBI Molecules]] being studied in the  [http://www.umass.edu/cbi/ University of Massachusetts Amherst Chemistry-Biology Interface Program] at UMass Amherst and on display at the [http://www.molecularplayground.org/ Molecular Playground].


'''Transcription and the human mitochondrial RNA polymerase '''
=== Transcription and the human mitochondrial RNA polymerase ===


The central dogma of biology in which genetic information is transferred from DNA to RNA, and subsequently into protein, is fundamental to life. A key player in this process is the DNA-dependent RNA polymerase. RNA polymerase produces RNA in the presence of a DNA template under tight control by the cell. In the Martin lab, our goal is to elucidate the energetics and thermodynamics of this complicated process. As a model, we use T7 bacteriophage RNA polymerase. This single-subunit polymerase can transcribe DNA without assistance from other proteins, making it an ideal model to understand transcription. Likewise, it is representative of all other known RNA polymerases in that it initiates at unique positions along the DNA, undergoes abortive cycling, transitions to a stable elongation complex, and terminates transcription at specific sequences.  
The central dogma of biology in which genetic information is transferred from DNA to RNA, and subsequently into protein, is fundamental to life. A key player in this process is the DNA-dependent RNA polymerase. RNA polymerase produces RNA in the presence of a DNA template under tight control by the cell. In the Martin lab, our goal is to elucidate the energetics and thermodynamics of this complicated process. As a model, we use T7 bacteriophage RNA polymerase (PDB:1QLN). This single-subunit polymerase can transcribe DNA without assistance from other proteins, making it an ideal model to understand transcription. Likewise, it is representative of all other known RNA polymerases in that it initiates at unique positions along the DNA, undergoes abortive cycling, transitions to a stable elongation complex, and terminates transcription at specific sequences.  


Related to the T7 RNA polymerase is the human mitochondrial RNA polymerase <scene name='User:Lynmarie_K_Thompson/Sandbox_1/Loadedfrompdb/4'> (PDB: 3SPA)</scene>. Recent crystallization of this enzyme by [http://www.ncbi.nlm.nih.gov/pubmed/21947009 Temiakov et al.] has revealed structural similarity between the two polymerases. The <font color =gray> C-terminal </font> <scene name='User:Lynmarie_K_Thompson/Sandbox_1/Loadedfrompdb/4'>domain</scene> contains the active site for RNA synthesis. The <font color=magenta> N-terminal </font> <scene name='User:Lynmarie_K_Thompson/Sandbox_1/Loadedfrompdb/4'> domain, </scene>(NTD) contains the promoter-binding domain (PBD) essential for DNA binding and start site specificity. Key structures of the NTD include the <font color=red> specificity </font> <scene name='User:Lynmarie_K_Thompson/Sandbox_1/Loadedfrompdb/4'>loop</scene>, <font color=orange> intercalating </font> <scene name='User:Lynmarie_K_Thompson/Sandbox_1/Loadedfrompdb/4'>loop</scene> , and <font color=blue> AT-recognition </font> <scene name='User:Lynmarie_K_Thompson/Sandbox_1/Loadedfrompdb/4'>loop</scene>.


One very big difference between the two polymerases is in the form of two transcription factors, TFB2M and TFAM. Human mtRNA polymerase requires these two proteins for binding and melting of promoter DNA. TFAM binds 15-40 base pairs upstream while TFB2M binds more proximal to the transcription start site (1-3). The requirement of these transcription factors coupled with the discrepancy of PBD orientation between T7 (Fig. 2; salmon) and human mtRNA polymerase (Fig. 2; grey) suggests a role in enzyme remodeling for the transcription factors, namely, TFB2M. By studying the structural changes and differences between the two polymerases, we can better under the energetics that governed transcription.


Initially transcribing complexes are relatively unstable through about the first 8-10 bases of transcription. During this time, short RNA transcripts are made and released in a process known as abortive cycling. Abortive cycling in particular, and promoter-proximal pausing more generally, plays key roles in gene regulation. Thus understanding the mechanism of initial transcription is key to understanding cellular regulation.
== References ==


#Ekstrand, M. I. Mitochondrial transcription factor A regulates mtDNA copy number in mammals. Hum. Mol. Genet. 13, 935-944 (2004). [http://www.ncbi.nlm.nih.gov/pubmed/15016765 Pubmed]
#Dairaghi, D. J., Shadel, G. S., Clayton, D. A. Human mitochondrial transcription factor A and promoter spacing integrity are required for transcription initiation. Biochim. Biophys. Acta 1271, 127-134 (1995). [http://www.ncbi.nlm.nih.gov/pubmed/7599198 Pubmed]
#Sologub, M., Litonin, D., Anikin, M., Mustaev, A., Temiakov, D. TFB2 is a transient component of the catalytic site of the human mitochondrial RNA polymerase. Cell 139, 934-944 (2009).ﾠ[http://www.ncbi.nlm.nih.gov/pubmed/19945377 Pubmed]


A bacterial chemotaxis receptor is an unusually long alpha-helical structure. The attractant molecule (the ligand) binds near the top of this picture and sends a signal across the membrane into the cell to control proteins that bind near the bottom. This is a model of the structure of the receptor based on experimental structures of pieces of related proteins.
== HELP! ==


{{Clear}}
I was unable to create Jmol scenes to illustrate structures of the human mitochondrial RNA polymerase due to a "java.security.AccessControlException: access denied" problem regardless of the computer or web browser I use.
<applet load='1wat' size='[450,338]' frame='true' align='right'
caption='Aspartate receptor ligand binding domain (1wat)' scene='User:Lynmarie_K_Thompson/Sandbox_1/Loadedfrompdb/4'/>


=== Ligand-binding domain ===
If there is anyone that may know a remedy for this problem, please contact kmathavan@mcb.umass.edu. Thank you!
 
 
 
The spinning protein (<scene name='User:Lynmarie_K_Thompson/Sandbox_1/Loadedfrompdb/4'>Initial view</scene>) ) is the ligand binding domain of the aspartate receptor with the aspartate ligand bound (LKT).
 
 
Molecular Playground banner: A bacterial chemotaxis receptor protein used by bacteria to "smell" their environment.
 
 
{{Clear}}
<applet load='2ho9' size='[450,338]' frame='true' align='right'
caption='E. coli chemotaxis adaptor protein CheW (2ho9)' scene='User:Shiela_M._Jones/Sandbox_1/Chew_suppressionmutants/1'/>
 
=== Chemotaxis adaptor protein CheW ===
 
 
CheW is a chemotaxis adaptor protein, and part of the tertiary complex formed by the chemotaxis receptor, histidine kinase protein CheA, and CheW. As an adaptor protein, CheW mediates the interaction between the chemotaxis receptor and CheA, and is necessary for the formation of kinase active complexes. CheW has been found to bind to the P5 domain of CheA through crystallographic studies.
 
At right, CheW is shown with suppression mutants (blue)that have been measured to decrease receptor binding and chemotaxis (SMJ).
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