Researchers explain how ubiquitin function is determined

University of Maryland researchers have explained, for the first time, why the linkage between the cell-regulating protein molecules called ubiquitins determines their function.

As reported in a paper in the June 10 issue of the journal Molecular Cell, Maryland biochemistry professor David Fushman and his research team used nuclear magnetic resonance to unveil the mechanism by which the lysine that joins the ubiquitins like pearls on a necklace determines which cellular operation the ubiquitin chains regulate.

“We confirmed that the linkage determines the way the ubiquitins are lined up, which, in turn, determines the shape of a ubiquitin chain and its signaling function,” said Fushman.

Better understanding of the workings of ubiquitins may help lead to new methods of drug therapy for the treatment of cancer or neurodegenerative diseases, such as Alzheimer’s and Parkinson’s.

Living up to their name, ubiquitin molecules are found everywhere in the life of the human cell, calling the shots for just about everything the cell does. “Ubiquitins are involved in regulating almost every aspect of a cell’s life,” says Fushman,” from mitosis to growth to communication to cell death.”

As important as ubiquitins are, however, scientists are only beginning to unlock the secrets of how these molecules go about their business. One of the things they do know is that ubiquitins can be connected to each other, like pearls on a necklace. When attached to a protein, the ubiquitin tag acts as a signal to the cell that determines the protein’s destiny. The “string” that holds the ubiquitins together in the chain is the amino acid lysine.

While ubiquitins are identical, the chains are not. They can be connected through one of seven lysines in ubiquitin. Different regulating signals are sent from different configurations of the connected ubiquitins. For instance, the chains connected with Lysine-63 (Lys-63) are involved in DNA repair and inflammatory response. The ubiquitins connected through Lysine-48 (Lys-48) target protein that needs to be discarded to its degradation in the ‘molecular shredder’ called proteasome.

“We knew that the chains are made of the same molecule, but they are giving different signals,” Fushman said, “Why? What makes these chains so different?”

Fushman’s lab had discovered earlier that two ubiquitins linked through Lys-48 can stick to each other in solution, while Lys-63 ubiquitins can’t. “One side of ubiquitin is hydrophobic, it ‘fears’ water,” says Fushman. “These molecules try to hide that side by sticking to each other, like Velcro, forming what we call a hydrophobic contact.”

“We knew that the Lys-48 linkage allows the units to bend the chain and make these contacts, or have a ‘compact conformation.’ The Lys-63 chain, on the other hand, is straight-- the ubiquitins cannot make the hydrophobic contact.

“Our hypothesis was that linkage through different lysines makes the chains do different things, but we didn’t have the structural details.”

Hoping to discover those structural details, Fushman, with doctoral student Ranjani Varadan and postdoctoral fellow Michael Assfalg, concentrated on the Lys-48 chain. They introduced a ubiquitin-associated domain (UBA), a piece of a protein Rad23a, which helps the proteasome recognize the ubiquitin chains that are delivering proteins for degradation.

Rad23a has been shown by Fushman’s collaborators Cecile Pickart and Shahri Raasi of Johns Hopkins to prefer Lys-48-linked chains to Lyse-63 chains.

“We wanted to find out what is so different in the Lys-48 chain that Rad23a prefers it,” Fushman said.

It turns out that what Rad23a-UBA prefers was that it can nestle in the hydrophobic pocket that Lys-48 linked chains make. “Both sides of UBA domain are hydrophobic,” explains Fushman. “On the Lys-48 chain it could bind separately to the ubiquitins on either side, like a hamburger in a bun. The hydrophobic pocket makes it safe. It can’t do that on the Lys-63–linked chain.”

For the first time, Fushman says, “we have shown where the UBA domain binds ubiquitin chains and what the structure of the complex is. This helps us understand how the conformation of the ubiquitin chain determines its function. We can now look at different chains formed using other links.”

Co-authors on the paper include Ranjani Varadan and Michael Assfalg, of the University of Maryland, and Cecile Pickart and Shahri Raasi of Johns Hopkins.


The opinions expressed here are the views of the writer and do not necessarily reflect the views and opinions of News Medical.
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