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Structure of human mitochondrial RNA polymerase elongation complex.

Schwinghammer K, Cheung AC, Morozov YI, Agaronyan K, Temiakov D, Cramer P - Nat. Struct. Mol. Biol. (2013)

Bottom Line: Whereas T7 RNAP refolds during the transition from initiation to elongation, mtRNAP adopts an intermediary conformation that is capable of elongation without refolding.The intercalating hairpin that melts DNA during T7 RNAP initiation separates RNA from DNA during mtRNAP elongation.Newly synthesized RNA exits toward the pentatricopeptide repeat (PPR) domain, a unique feature of mtRNAP with conserved RNA-recognition motifs.

View Article: PubMed Central - PubMed

Affiliation: Gene Center, Department of Biochemistry, Ludwig-Maximilians-Universität München, Munich, Germany.

ABSTRACT
Here we report the crystal structure of the human mitochondrial RNA polymerase (mtRNAP) transcription elongation complex, determined at 2.65-Å resolution. The structure reveals a 9-bp hybrid formed between the DNA template and the RNA transcript and one turn of DNA both upstream and downstream of the hybrid. Comparisons with the distantly related RNA polymerase (RNAP) from bacteriophage T7 indicates conserved mechanisms for substrate binding and nucleotide incorporation but also strong mechanistic differences. Whereas T7 RNAP refolds during the transition from initiation to elongation, mtRNAP adopts an intermediary conformation that is capable of elongation without refolding. The intercalating hairpin that melts DNA during T7 RNAP initiation separates RNA from DNA during mtRNAP elongation. Newly synthesized RNA exits toward the pentatricopeptide repeat (PPR) domain, a unique feature of mtRNAP with conserved RNA-recognition motifs.

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Lack of NTD refolding upon mtRNAP elongation observed in the crystal structure(a–c) Structures of the NTDs of T7 RNAP and mtRNAP. The NTD of T7 RNAP (a) is refolded in the elongation complex (PDB 1MSW34), whereas the NTD of mtRNAP (b) is not, and resembles the NTD in the T7 intermediate (PDB 3E2E15) (c). Helices are depicted as cylinders and nucleic acids as ribbons with sticks for protruding bases.(d) The FG loop of T7 RNAP (PDB 1QLN4, pale cyan) protrudes into the hybrid-binding site but is shorter and positioned differently in mtRNAP (silver).
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Figure 5: Lack of NTD refolding upon mtRNAP elongation observed in the crystal structure(a–c) Structures of the NTDs of T7 RNAP and mtRNAP. The NTD of T7 RNAP (a) is refolded in the elongation complex (PDB 1MSW34), whereas the NTD of mtRNAP (b) is not, and resembles the NTD in the T7 intermediate (PDB 3E2E15) (c). Helices are depicted as cylinders and nucleic acids as ribbons with sticks for protruding bases.(d) The FG loop of T7 RNAP (PDB 1QLN4, pale cyan) protrudes into the hybrid-binding site but is shorter and positioned differently in mtRNAP (silver).

Mentions: To initiate transcription, T7 RNAP binds promoter DNA with its NTD7,22. The NTD then refolds during the transition from an initiation complex4 to an elongation complex16 via an intermediary state15. In contrast, the NTD of mtRNAP does not refold during the initiation–elongation transition (Fig. 5). The NTD fold observed in our mtRNAP elongation complex structure differs from that in T7 RNAP elongation complexes, but resembles that in the T7 initiation–elongation intermediate, and is partially related to that in the T7 initiation complex (Figs. 5a–c and Supplementary Table 2 online).


Structure of human mitochondrial RNA polymerase elongation complex.

Schwinghammer K, Cheung AC, Morozov YI, Agaronyan K, Temiakov D, Cramer P - Nat. Struct. Mol. Biol. (2013)

Lack of NTD refolding upon mtRNAP elongation observed in the crystal structure(a–c) Structures of the NTDs of T7 RNAP and mtRNAP. The NTD of T7 RNAP (a) is refolded in the elongation complex (PDB 1MSW34), whereas the NTD of mtRNAP (b) is not, and resembles the NTD in the T7 intermediate (PDB 3E2E15) (c). Helices are depicted as cylinders and nucleic acids as ribbons with sticks for protruding bases.(d) The FG loop of T7 RNAP (PDB 1QLN4, pale cyan) protrudes into the hybrid-binding site but is shorter and positioned differently in mtRNAP (silver).
© Copyright Policy
Related In: Results  -  Collection

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Show All Figures
getmorefigures.php?uid=PMC4321815&req=5

Figure 5: Lack of NTD refolding upon mtRNAP elongation observed in the crystal structure(a–c) Structures of the NTDs of T7 RNAP and mtRNAP. The NTD of T7 RNAP (a) is refolded in the elongation complex (PDB 1MSW34), whereas the NTD of mtRNAP (b) is not, and resembles the NTD in the T7 intermediate (PDB 3E2E15) (c). Helices are depicted as cylinders and nucleic acids as ribbons with sticks for protruding bases.(d) The FG loop of T7 RNAP (PDB 1QLN4, pale cyan) protrudes into the hybrid-binding site but is shorter and positioned differently in mtRNAP (silver).
Mentions: To initiate transcription, T7 RNAP binds promoter DNA with its NTD7,22. The NTD then refolds during the transition from an initiation complex4 to an elongation complex16 via an intermediary state15. In contrast, the NTD of mtRNAP does not refold during the initiation–elongation transition (Fig. 5). The NTD fold observed in our mtRNAP elongation complex structure differs from that in T7 RNAP elongation complexes, but resembles that in the T7 initiation–elongation intermediate, and is partially related to that in the T7 initiation complex (Figs. 5a–c and Supplementary Table 2 online).

Bottom Line: Whereas T7 RNAP refolds during the transition from initiation to elongation, mtRNAP adopts an intermediary conformation that is capable of elongation without refolding.The intercalating hairpin that melts DNA during T7 RNAP initiation separates RNA from DNA during mtRNAP elongation.Newly synthesized RNA exits toward the pentatricopeptide repeat (PPR) domain, a unique feature of mtRNAP with conserved RNA-recognition motifs.

View Article: PubMed Central - PubMed

Affiliation: Gene Center, Department of Biochemistry, Ludwig-Maximilians-Universität München, Munich, Germany.

ABSTRACT
Here we report the crystal structure of the human mitochondrial RNA polymerase (mtRNAP) transcription elongation complex, determined at 2.65-Å resolution. The structure reveals a 9-bp hybrid formed between the DNA template and the RNA transcript and one turn of DNA both upstream and downstream of the hybrid. Comparisons with the distantly related RNA polymerase (RNAP) from bacteriophage T7 indicates conserved mechanisms for substrate binding and nucleotide incorporation but also strong mechanistic differences. Whereas T7 RNAP refolds during the transition from initiation to elongation, mtRNAP adopts an intermediary conformation that is capable of elongation without refolding. The intercalating hairpin that melts DNA during T7 RNAP initiation separates RNA from DNA during mtRNAP elongation. Newly synthesized RNA exits toward the pentatricopeptide repeat (PPR) domain, a unique feature of mtRNAP with conserved RNA-recognition motifs.

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Related in: MedlinePlus