User:Momodou L. Jammeh/Sandbox 1

From Proteopedia

proteopedia linkproteopedia link

Revision as of 11:19, 29 April 2011 (edit) Momodou L. Jammeh (Talk | contribs) ← Previous diff		Revision as of 11:35, 29 April 2011 (edit) (undo) Momodou L. Jammeh (Talk | contribs) Next diff →
Line 19:		Line 19:
	<scene name='User:Momodou_L._Jammeh/Sandbox_1/Rho_p-loop/2'>C-terminus structures</scene>		<scene name='User:Momodou_L._Jammeh/Sandbox_1/Rho_p-loop/2'>C-terminus structures</scene>
		+
		+	<scene name='User:Momodou_L._Jammeh/Sandbox_1/Subunit-c-terminal/1'>C_terminal</scene>
	----		----
	===MECHANISM===		===MECHANISM===

---

Revision as of 11:35, 29 April 2011

---

Contents 1 RHO TERMINATION FACTOR 1.1 INTRODUCTION 1.2 STRUCTURE 1.3 MECHANISM 1.4 INHIBITION

RHO TERMINATION FACTOR

---

INTRODUCTION

Transcription is the process of mRNA synthesis by RNA polymerase, an enzyme that uses a strand of DNA as a template for ribonucloetide addition. During this process, RNA polymerase encounters several regulatory proteins, called transcription factors, that effect transcription in various ways. One of the major regulatory activities of transcription regulators is terminating the process at specific sites in the transcription unit. In E. coli and prokaryotes in general, transcription is terminated through rho dependent and independent mechanisms. Transcription termination without rho occurs through RNA polymerase's intrinsic ability to terminate transcription in response to certain, limited sequences in the template DNA strand. Rho dependent termination requires a helicase protein called rho factor which belongs in the helicase class of enzymes. Helicases are motor proteins that function through directional movement on the nucleic acid phosphodiester backbone and can dissociate bound DNA-DNA, RNA-DNA or RNA-RNA strands. Rho specifically terminates transcription by separating the the new RNA strand annealed to the DNA strand being transcribed. As pictured on the upper right corner, it is a hexameric ring-shaped helicase that uses energy derived from its ATPase mechanism (hydrolysis of ATP into ADP and an organic phosphate) to drive its movement along the newly formed RNA molecule toward the elongation complex to be dissociated. While rho is only requirement for the termination process, additional elongation factors such as NusA and NusG have been shown to enhance the process. NusA reduces the elongation rate allowing rho to catch up with RNA polymerase despite its high processivity while NusG enhances the rate of RNA release by coupling rho to the elongation complex. Since its discovery in 1969, studies on rho have been essential in understanding how external factors regulation transcription termination and its structure provides insight into how several stable hexameric helicases load onto their nucleic acid substrates.

---

STRUCTURE

spinqualitylabelsall models Rho factor in its hexameric form with its six identical subunits forming a helicase and an RNA molecule right through the center Show:Asymmetric Unit Biological Assembly Drag the structure with the mouse to rotate   Export Animated Image

Rho factor is a hexamer of six identical subunits that interact to form a closed, dynamic structure called a It is made up of a single polypeptide chain of about 419 amino acid residues which comprise the and conformations that make up the (single subunit). Each subunit has an (residues in blue and light blue) and (residues in red and orange) domain with the former on the periphery of the subunits and the latter, which is also the main catalytic site on the central, upper portion of the hexamer. The domain of the rho (A chain) consists of amino acid residues 1 - 130 and serves as the primary binding site for the C-rich of the nascent RNA transcript. It has an formed from five polypeptide strands that comprise the two anti parallel sheets that are connected by four loops. The barrel is closed by hydrogen bonds between the third and fifth strands and forms the oligonucleotide/oligosaccharide binding (OB) fold of rho. The is a folding motif that is common among oligonucleotide or oligosaccharide binding proteins. Although OB folds have been found in several other proteins such as staphylococcal nuclease and anti codon binding regions of asp-tRNA synthetase, the geometry of the OB fold (five strand beta-barrel of two antiparallel sheets, otherwise known as the greek key motif) is the same for all proteins but their is no sequence similarity. Therefore, the binding of these proteins to their oligonucleotide/oligosaccharide substrates depends on fold geometry and topology rather than on the protein sequences. Because of the presence of the in the N-terminal domain of rho, it is evident that this sub-domain constitutes the primary binding site for the RNA transcript as it loads on to rho. As stated earlier, the is the main catalytic site of rho and has several structural motifs to that effect. The major catalytic process that occurs in this domain is the hydrolysis of ATP and thus contains the Walker A and Walker B signature motifs that are common to such ATPases. Here, an sits in the ATP binding pocket of adjoining A and B subunits of rho. The Walker A motif is the site for ATP binding and contains the which has the consensus amino acid sequence GXXXXGK(T/S), with the K denoting the which is crucial for nucleotide binding. In addition to LYS, interact with bound ATP. The coordinates the γ- and β-phosphates of nucleotides. The Walker B motif which is characterized by the and facilitates mRNA translocation and unwinding and thus comprises the secondary binding site of the .

Rho is assemble through subunit-subunit interactions between both the amino- and carboxyl terminus domains of the six subunits. Adjacent N-terminal domains make contact through α2/α3 connector loop, while the C-terminus interactions occur through the α11 which packs against the β7/α8 and β8/α9 junctions of the neighboring subunit. Although, extensive reference has been made to existence of rho as a closed ring, it is also thought to split open to yield the "" conformation. An unusual conformation considering the fact that almost all fully crystallized, full length structures of rho are in closed ring form.

---

MECHANISM

The termination of transcription by rho requires the binding of nascent RNA by the hexamer which functions as an RNA-activated ATPase helicase. This molecular motor is driven by ATP hydrolysis which occurs in the main catalytic site in the C-terminal domain of rho monomers. Rho makes contact with its substrate, nascent RNA transcript, through its OB fold which serves as the primary RNA binding site. The OB fold fold recognizes and binds to the cytosine-rich region of a transcript called the rho utilization (rut) sequence of 40 or more bases. Thus, the N-terminus domain makes first contact with RNA and the loading process begins.

---

INHIBITION

---