Forum for Science, Industry and Business

Sponsored by:     3M 
Search our Site:

 

Nobel Laureate finds 'elegant' explanation for DNA transcribing enzyme's high fidelity

01.12.2006
Last month, Roger Kornberg of Stanford University won the Nobel Prize in Chemistry for his efforts to unravel the molecular basis of eukaryotic transcription, in which enzymes give “voice” to DNA by copying it into the RNA molecules that serve as templates for protein in organisms from yeast to humans.

Now, Kornberg and his colleagues report in the December 1, 2006 issue of the journal Cell, published by Cell Press, new structures that reveal another critical piece of the puzzle: how the so-called polymerase II enzyme discriminates among potential RNA building blocks to ensure the characteristic accuracy of the process.

The researchers found that a portion of the enzyme known as the trigger loop acts like a “trap door,” swinging beneath a matching nucleoside triphosphate (NTP) building block, to close off the active center and form an extensive network of interactions with the NTP and other parts of the enzyme. Those interactions leave another side chain in the trigger loop precisely positioned, such that it may literally “trigger” the formation of the chemical bonds that link components of the growing RNA chain together. If the NTP is even slightly misaligned, Kornberg said, those critical interactions fail.

The trigger loop mechanism therefore couples NTP recognition and catalysis, ensuring the fidelity of transcription, they reported.

... more about:
»DNA »Kornberg »NTP »Polymerase »RNA »explanation »transcribing

“Of all revelations from the structure [of the transcription machinery] since it was first solved, this is perhaps the most fundamental since it gets at the underlying mechanisms,” Kornberg said. “It’s long known that the enzyme operates with high fidelity—selecting the correct base and sugar—but it’s been a mystery how that is accomplished.”

These findings offer “an unexpected and elegant explanation that’s both beautiful and simple, as nature invariably proves to be.”

The fundamental mechanism of transcription is conserved among cellular RNA polymerases, the researchers explained. Common features include an unwound region of about 15 base pairs of the DNA with some eight residues of the RNA transcript hybridized with the DNA in the center of the “transcription bubble.” The enzyme polymerases involved are capable of moving both forward and backward on the DNA. Forward movement is favored by the binding of NTPs, while backtracking occurs especially when the enzyme encounters an impediment, such as damaged DNA.

Kornberg’s group captured the first picture of the polymerase II transcribing complex by X-ray crystallography in 2001. Those structures revealed the complex with a nucleotide still in the enzyme’s addition site, just after it had been added to the RNA transcript.

Later X-ray structures revealed the transcribing complex with the addition site available for entry of a matched NTP. Those crystals uncovered a second NTP-binding site on the transcribing enzyme, dubbed the entry site. While all NTPs can bind the entry site, only an NTP matched for base-pairing with the DNA template binds the addition site for attachment to the growing RNA strand, Kornberg said.

Yet the question of how the enzyme achieves such a high degree of discrimination between matched and mismatched NTPs remained unanswered.

The chemical attraction alone between RNA bases—adenine, cytosine, guanine, and uracil—and their complementary bases on the DNA template strand is far from sufficient to account for the incredible selectivity of polymerase II, Kornberg said. And the scientists didn’t know either how the polymerase avoids substituting the NTPs that constitute DNA for the correct RNA building blocks, molecules that differ by only one oxygen atom.

In search of an explanation in the current study, the researchers screened hundreds of crystals to achieve higher data quality and resolution than ever before.

“In the course of the work, we saw something that had never been noticed before— additional protein density beneath the matching nucleotide,” Kornberg said.

The team traced that protein density back to a portion of the polymerase II enzyme: the trigger loop.

“Of the 14 crystal structures now reported in which the trigger loop was observed, only in two is it seen in that location, directly beneath the NTP,” Kornberg said. Those were the only two crystals in which the NTP was correctly matched to the DNA template, evidence of the trigger loop’s “clear relationship to NTP selection.”

Further study revealed that, when a matching NTP reaches the addition site, the trigger loop swings from its usual position some distance away until it rests parallel to the NTP. It then forms a network of interactions—some 20 to 30 in all—with components of the NTP, a process that serves to “recognize all features of the NTP in the addition site and detect its precise location,” the researchers reported.

“The specificity is a result of the alignment with the NTP that is critically dependent upon the base, sugar, phosphate and location when the trigger loop swings into position,” Kornberg said. “If it is misaligned even slightly, that set of contacts cannot occur.”

As a consequence of that alignment, to angstrom (a unit of length equal to one hundred millionth of a centimeter) precision, a histidine side chain of the trigger loop rests on the ß phosphate, the chemical constituent that must have its bond broken in order for the NTP to join the RNA chain through the formation of a phosphodiester bond, Kornberg said. The finding suggested the side chain acts as a trigger for bond formation.

The whole decision-making process occurs extremely rapidly, he added, on the order of picoseconds. A picosecond is one trillionth of a second.

“The basis for the extraordinary specificity with which RNA polymerases transcribe DNA lies in a structural element termed the trigger loop, which makes both direct and indirect contact with all features of the nucleotide in the polymerase active center,” the researchers concluded.

Heidi Hardman | EurekAlert!
Further information:
http://www.cell.com

Further reports about: DNA Kornberg NTP Polymerase RNA explanation transcribing

More articles from Life Sciences:

nachricht How brains surrender to sleep
23.06.2017 | IMP - Forschungsinstitut für Molekulare Pathologie GmbH

nachricht A new technique isolates neuronal activity during memory consolidation
22.06.2017 | Spanish National Research Council (CSIC)

All articles from Life Sciences >>>

The most recent press releases about innovation >>>

Die letzten 5 Focus-News des innovations-reports im Überblick:

Im Focus: Can we see monkeys from space? Emerging technologies to map biodiversity

An international team of scientists has proposed a new multi-disciplinary approach in which an array of new technologies will allow us to map biodiversity and the risks that wildlife is facing at the scale of whole landscapes. The findings are published in Nature Ecology and Evolution. This international research is led by the Kunming Institute of Zoology from China, University of East Anglia, University of Leicester and the Leibniz Institute for Zoo and Wildlife Research.

Using a combination of satellite and ground data, the team proposes that it is now possible to map biodiversity with an accuracy that has not been previously...

Im Focus: Climate satellite: Tracking methane with robust laser technology

Heatwaves in the Arctic, longer periods of vegetation in Europe, severe floods in West Africa – starting in 2021, scientists want to explore the emissions of the greenhouse gas methane with the German-French satellite MERLIN. This is made possible by a new robust laser system of the Fraunhofer Institute for Laser Technology ILT in Aachen, which achieves unprecedented measurement accuracy.

Methane is primarily the result of the decomposition of organic matter. The gas has a 25 times greater warming potential than carbon dioxide, but is not as...

Im Focus: How protons move through a fuel cell

Hydrogen is regarded as the energy source of the future: It is produced with solar power and can be used to generate heat and electricity in fuel cells. Empa researchers have now succeeded in decoding the movement of hydrogen ions in crystals – a key step towards more efficient energy conversion in the hydrogen industry of tomorrow.

As charge carriers, electrons and ions play the leading role in electrochemical energy storage devices and converters such as batteries and fuel cells. Proton...

Im Focus: A unique data centre for cosmological simulations

Scientists from the Excellence Cluster Universe at the Ludwig-Maximilians-Universität Munich have establised "Cosmowebportal", a unique data centre for cosmological simulations located at the Leibniz Supercomputing Centre (LRZ) of the Bavarian Academy of Sciences. The complete results of a series of large hydrodynamical cosmological simulations are available, with data volumes typically exceeding several hundred terabytes. Scientists worldwide can interactively explore these complex simulations via a web interface and directly access the results.

With current telescopes, scientists can observe our Universe’s galaxies and galaxy clusters and their distribution along an invisible cosmic web. From the...

Im Focus: Scientists develop molecular thermometer for contactless measurement using infrared light

Temperature measurements possible even on the smallest scale / Molecular ruby for use in material sciences, biology, and medicine

Chemists at Johannes Gutenberg University Mainz (JGU) in cooperation with researchers of the German Federal Institute for Materials Research and Testing (BAM)...

All Focus news of the innovation-report >>>

Anzeige

Anzeige

Event News

Plants are networkers

19.06.2017 | Event News

Digital Survival Training for Executives

13.06.2017 | Event News

Global Learning Council Summit 2017

13.06.2017 | Event News

 
Latest News

Quantum thermometer or optical refrigerator?

23.06.2017 | Physics and Astronomy

A 100-year-old physics problem has been solved at EPFL

23.06.2017 | Physics and Astronomy

Equipping form with function

23.06.2017 | Information Technology

VideoLinks
B2B-VideoLinks
More VideoLinks >>>