Important new information on one of the most critical protein machines in living cells has been reported by a team of researchers with the U.S. Department of Energy’s Lawrence Berkeley National Laboratory (Berkeley Lab) and the University of California (UC) Berkeley.
The researchers have provided the most detailed look ever at the “regulatory particle” used by the protein machines known as proteasomes to identify and degrade proteins that have been marked for destruction. The activities controlled by this regulatory particle are critical to the quality control of cellular proteins, as well as a broad range of vital biochemical processes, including transcription, DNA repair and the immune defense system.
“Using electron microscopy and a revolutionary new system for protein expression, we have determined at a subnanometer scale the complete architecture, including the relative positions of all its protein components, of the proteasome regulatory particle,” says biophysicist Eva Nogales, the research team’s co-principal investigator. “This provides a structural basis for the ability of the proteasome to recognize and degrade unwanted proteins and thereby regulate the amount of any one type of protein that is present in the cell.”
Says the team’s other co-principal investigator and corresponding author, biochemist Andreas Martin, “While the biochemical function of many of the proteasome components have been determined, and some subnanometer structures have been identified, it was unclear before now which component goes where and which components interact with one another. Now we have a much better understanding as to how the proteasome machinery works to control cellular processes and this opens the possibility of manipulating proteasome activity for the treatment of cancer and other diseases.”
Nogales, who holds appointments with Berkeley Lab, UC Berkeley, and the Howard Hughes Medical Institute, and Martin, who holds appointments with UC Berkeley and the QB3 Institute, are the senior authors of a paper describing this work in the journal Nature. The paper is titled “Complete subunit architecture of the proteasome regulatory particle.” Other co-authors were Gabriel Lander, Eric Estrin, Mary Matyskiela and Charlene Bashore.
At any given moment, a human cell typically contains about 100,000 different proteins, with certain proteins being manufactured and others being discarded as needed for the cell’s continued prosperity. Unwanted proteins are tagged with a “kiss-of-death” label in the form of a polypeptide called “ubiquitin.” A protein marked with ubiquitin is delivered to any one of the some 30,000 proteasomes in the cell – barrel-shaped complexes which act as waste disposal units that rapidly break-down or degrade the protein. The 2004 Nobel Prize in chemistry was awarded to a trio of scientists who first described the proteasome process, but a lack of structural information has limited the scientific understanding of the mechanics behind this process.
Nogales, an expert on electron microscopy and image analysis, and Martin, who developed the new protein expression system used in this work, combined the expertise of their respective research groups to study the proteasome regulatory particle in yeast. The particle features 19 sub-units that are organized into two sub-complexes, a “lid” and a “base.” The lid contains the regulatory elements that identify the ubiquitin tag marking a protein for destruction, and the base features a hexameric ring that pulls the tagged protein inside the chamber of the proteasome barrel where it is degraded.
“The lid consists of nine non-ATPase proteins including ubiquitin receptors that accept properly tagged proteins but prevent a protein not marked for degradation from engaging with the proteasome,” Nogales says. “Since degradation is irreversible, it is critical that only ubiquitin-tagged proteins engage the proteasome. Interestingly, the ubiquitin tag has to be removed before the protein can be translocated into the proteasome’s destruction chamber, so the lid also contains de-ubiquitination enzymes that remove the tags after the protein has engaged with the proteasome.”
The proteasome regulatory particle’s base contains six distinct AAA+ ATPases that form the hetero-hexameric ring, which serves as the molecular motor of the proteasome.
“We predict that the ATPases use the energy of ATP binding and hydrolysis to exert a pulling force on engaged proteins, unfolding and translocating them through a narrow central pore and into the degradation chamber,” Martin says. “The steps in the proteasome process – from protein recognition to de-ubiquitination and degradation have to be very highly coordinated in time and space. Locating all of these components and identifying their relative orientations has been very telling about how the processes are coordinated with each other.”
Nogales credits the protein expression system developed by Martin and his research group, in which proteins are expressed and assembled in bacteria, as being critical to the success of this research.
“Until now researchers had to work with purified protein complexes from the cell, which could not be manipulated or modified in any way,” she says. “Andy Martin’s new heterologous expression system allows for the manipulation and dissection of protein functions. For our studies it was crucial to generate lid sub-complexes that had one marker at a time in each of the subunits so that we could determine the position of each protein within the lid. With this new system we generated truncations, deletions and fusion constructs that were used to localize individual subunits and delineate their boundaries within the lid.”
This research was supported by funds from UC Berkeley, Berkeley Lab, the National Institutes of Health, the Searle Scholars Program, the Damon Runyon Cancer Research Foundation, the American Cancer Society, the National Science Foundation and the Howard Hughes Medical Institute.
Lawrence Berkeley National Laboratory addresses the world’s most urgent scientific challenges by advancing sustainable energy, protecting human health, creating new materials, and revealing the origin and fate of the universe. Founded in 1931, Berkeley Lab’s scientific expertise has been recognized with 13 Nobel prizes. The University of California manages Berkeley Lab for the U.S. Department of Energy’s Office of Science. For more, visit www.lbl.gov.
For more information about Eva Nogales and her research group see http://cryoem.berkeley.edu/
For more information about Andreas Martin and his research group see http://mcb.berkeley.edu/labs/martin/amartin/Home.html
Lynn Yarris | EurekAlert!
Further reports about: > Cancer > Gates Foundation > Living Lakes-Konferenz > Medical Wellness > Nobel Prize > Nogales > Waste-Disposal > cellular process > cellular protein > chemical process > electron microscopy > living cell > methanol fuel cells > principal investigator > protein expression > protein function > specimen processing
Modern genetic sequencing tools give clearer picture of how corals are related
17.08.2017 | University of Washington
The irresistible fragrance of dying vinegar flies
16.08.2017 | Max-Planck-Institut für chemische Ökologie
Whether you call it effervescent, fizzy, or sparkling, carbonated water is making a comeback as a beverage. Aside from quenching thirst, researchers at the University of Illinois at Urbana-Champaign have discovered a new use for these "bubbly" concoctions that will have major impact on the manufacturer of the world's thinnest, flattest, and one most useful materials -- graphene.
As graphene's popularity grows as an advanced "wonder" material, the speed and quality at which it can be manufactured will be paramount. With that in mind,...
Physicists at the University of Bonn have managed to create optical hollows and more complex patterns into which the light of a Bose-Einstein condensate flows. The creation of such highly low-loss structures for light is a prerequisite for complex light circuits, such as for quantum information processing for a new generation of computers. The researchers are now presenting their results in the journal Nature Photonics.
Light particles (photons) occur as tiny, indivisible portions. Many thousands of these light portions can be merged to form a single super-photon if they are...
For the first time, scientists have shown that circular RNA is linked to brain function. When a RNA molecule called Cdr1as was deleted from the genome of mice, the animals had problems filtering out unnecessary information – like patients suffering from neuropsychiatric disorders.
While hundreds of circular RNAs (circRNAs) are abundant in mammalian brains, one big question has remained unanswered: What are they actually good for? In the...
An experimental small satellite has successfully collected and delivered data on a key measurement for predicting changes in Earth's climate.
The Radiometer Assessment using Vertically Aligned Nanotubes (RAVAN) CubeSat was launched into low-Earth orbit on Nov. 11, 2016, in order to test new...
A study led by scientists of the Max Planck Institute for the Structure and Dynamics of Matter (MPSD) at the Center for Free-Electron Laser Science in Hamburg presents evidence of the coexistence of superconductivity and “charge-density-waves” in compounds of the poorly-studied family of bismuthates. This observation opens up new perspectives for a deeper understanding of the phenomenon of high-temperature superconductivity, a topic which is at the core of condensed matter research since more than 30 years. The paper by Nicoletti et al has been published in the PNAS.
Since the beginning of the 20th century, superconductivity had been observed in some metals at temperatures only a few degrees above the absolute zero (minus...
16.08.2017 | Event News
04.08.2017 | Event News
26.07.2017 | Event News
17.08.2017 | Physics and Astronomy
17.08.2017 | Materials Sciences
17.08.2017 | Materials Sciences