Correctly dismantling a structure can be as challenging as assembling it. The architecture of the yeast proteasome reveals this enzyme’s intricate machinery for protein degradation.

Thus begins an article in Nature about the cell’s shredder-recycler, the proteasome.¹ This large, barrel-shaped molecular machine has a flip-top lid like those trash cans with the foot pedal, only this one is much more elaborate: it validates the trash, pulls it in with a motor, and shreds it inside. New high-resolution images of the lid portion by Lander et al., also in Nature, reveal the workings of this “intricate architecture” as never before.²

This “massive proteolytic machine” (1.5 million atomic mass units, or 1.5 Mega-daltons) is composed of numerous protein subunits arranged in functional complexes, and is capable of degrading a wide variety of protein types. The barrel portion of the machine, where protein degradation occurs, had already been described in more detail. Inside the barrel (a stack of four rings with a cavity in the middle, composed of 28 protein parts), active sites on the inside walls cleave polypeptide chains into short segments about 7-9 amino acid units long, which can be reused by the cell directly, or further degraded into individual amino acids for recycling.

Obviously, this dangerous interior must be protected, lest it run amok like a chainsaw murderer. That’s why an elaborate lid structure, composed of 19 more protein parts, guards the entry gate and checks the credentials of each protein that enters.

To be validated, a target protein must have been previously tagged for destruction by other molecular machines. The tag is a ubiquitous protein called (appropriately) ubiquitin. Found in all eukaryotic cells, ubiquitin is encoded by redundant genes to ensure an ample supply is available at all times. Before being used as a tag, ubiquitin must be activated by additional enzymes through a sequence of checks and balances. Then, the tag is fastened onto a “tail” of the target protein, an unfolded portion long enough to fit into the proteasome chamber. Other enzymes then attach additional ubiquitin tags. The proteasome requires four ubiquitin tags to allow entry. Once tagged, the doomed protein is conveyed to the proteasome by processes that remain to be understood.

The entry into the proteasome is governed by two main complexes, the lid and the base, that lock together. The lid, called a “regulatory particle,” is much more than a mere “particle.” It is a complex of specific proteins tasked with recognizing the ubiquitin tags, removing them, initiating the unfolding of the target protein, and starting its descent into the barrel. Just underneath the lid, the base is a complex of six proteins forming a ring over the barrel opening. It grabs validated proteins and threads them into the chamber.

That much was known. For years, biochemists have been eager to understand how the validation, de-tagging, unfolding and initiation processes work. Here are the exciting new finds that Lander et al.² have revealed in sub-nanometer resolution, about the regulatory particle (“lid”) and base:

1. When isolated from the proteasome, the lid changes its structure to prevent exposure to its de-tagging machinery.
2. The lid fits the base at an angle, and makes extensive contacts with both the base and the barrel core.
3. When the lid docks to the base, parts move to grip the base and expose the active sites.
4. Two proteins on opposite sides of the lid serve as docking points for the ubiquitin tags. Although only one is necessary to initiate the degradation process, the two can work independently or together.
5. The distance between the tag-docking protein and the tag-cleaving protein ensures that four ubiquitin tags are present.
6. The “tail” of the target protein is inserted into the core.
7. Once the tags have been validated, another protein cleaves the tag from the tail.
8. Another protein off to the side separates the four tags into individual ubiquitin molecules.
9. The six proteins in the base are arranged like a spiral staircase. This means that either the proteins use a rotary mechanism to thread the polypeptide into the barrel, or remain statically in place as their individual moving parts grab and “walk” the polypeptide down the stairs.

A little imagination can help put this all into perspective. Think of a city’s recycling program. Let’s say the project wants to shred origami objects to reuse the paper. First, the objects have to identified and separated from the ones to be kept. They are tagged and sent to the recycling machine. The machine reads the tag and validates it, takes the tag off, unfolds the origami object, and sends it into shredder. The shredder grabs the material and cuts it into strips, which are then sent to another department for reconstitution. The tags are also recycled.

How many workers would it take to accomplish this? Each one has to know its job, know the process, and follow it faithfully; plus, a foreman has to supervise the work.

In the cell, all this is done through coding (both genetic and epigenetic) and automated processing. Proteasomes move throughout the cytoplasm and the nucleus, carrying on their vital work every day inside your body, identifying and selecting proteins to degrade, keeping the cell free of damaged or misfolded proteins, and removing proteins whose work is done. The proteasome can even activate certain proteins that require a “snip” to start working. These intricate machines help keep the immune system functioning and are ever-present to respond to stress. Failure of these systems can cause serious diseases, like Parkinson’s and Alzheimer’s, or death.

The authors of the original paper said nothing about evolution. Indeed, they ended just shy of a song of praise: “The intricate architecture of the proteasome highlights the complex requirements for this proteolytic machine, which must accommodate and specifically regulate a highly diverse set of substrates in the eukaryotic cell.”

Geng Tian and Daniel Finley, though, in their summary of the paper in Nature,¹ couldn’t resist sprinkling a little Darwin-brand sneeze powder on stage. “The eukaryotic proteasome seems to have evolved from a protease known as PAN (or something comparable to this enzyme), which is found in microorganisms called archaea,” they suggested. Then they produced a colorful diagram called “The evolution of proteases,” showing trypsin, PAN, and the full-fledged proteasome, as if to mimic those outworn icons of evolution, the horse series and monkey-to-man parade.

If all else fails, say it with feeling: “This dramatic evolutionary elaboration of the protein-degrading machinery is reflected in the fact that the proteasome has assumed regulatory functions in virtually all aspects of eukaryotic cell biology.” Before we can stop sneezing, there then comes the encore: “The evolution of ubiquitin tagging also coincided with a transformation of the proteasome’s structure.”

Did they show how mutations and unguided processes produced this highly complex machine, let alone the other systems it interacts with? No; they just assumed it. “The evolution of” begs the question. Comparing three functional designs of varying complexity doesn’t prove they evolved any more than showing a scissors, a shredder and a recycling system proves they evolved from each other by undirected processes.

“The evolution of” is a useless if ubiquitous phrase that needs to be tossed into the recycler.

Photo credit: maury.mccown, Flickr.

References Cited
1. Geng Tian and Daniel Finley, “Cell biology: Destruction deconstructed,” Nature 482, (09 February 2012), pp. 170-171, doi:10.1038/482170a.
2. Lander et al., “Complete subunit architecture of the proteasome regulatory particle,” Nature 482, (09 February 2012), pp. 186-191, doi:10.1038/nature10774.