Gone in 60 (milli)seconds

Intracellular proteins have to be degraded, more or less at the same rate as new proteins are produced (or the cell would eventually burst). On the other hand, you can’t go about degrading proteins willy-nilly.  There are vast and complex systems for identifying proteins that should be destroyed, tagging them, and then moving them into a controlled destruction chamber.

The most important of these systems is the ubiquitin-proteasome degradation pathway.  Proteins that are destined for destruction are tagged with a chain of ubiquitin molecules.1  There are multiple steps in this pathway, in which ubiquitin is prepared for tagging, target proteins are identified, and ubiquitin is transferred from the activating components to the targeted protein.

Target proteins are destroyed when a chain of ubiquitin molecules (head to tail) are attached to them. An unanswered question has been how this works. Is the ubiquitin chain formed first, and then transferred to the target en bloc? Or are single ubiquitin transferred one at a time, sequentially, first to the target protein and then to the previously-attached ubiquitins?  The problem has been that the process goes so fast that it’s been hard to distinguish between the possibilities.

Now, in a gorgeous series of experiments, Pierce et al2 were able to watch ubiquitination happening over fractions of a second:

… we performed our single-encounter reactions on a quench flow apparatus that allowed us to take measurements on a timescale ranging from 10 ms to 30 s2

And the answer looks pretty clear: Ubiquitins are transferred sequentially, not en bloc.

Even at this timescale, though, they weren’t able to catch the very first event — the transfer of the first ubiquitin to the target.  That happens, apparently, in less than 10-20 milliseconds.  They also draw the conclusion that target tagging is critically dependent on the kinetics of ubiquitin chain elongation (as you’d expect) which are governed by ubiquitin off-rates, and this mode of regulation is probably a billion years old.

Pierce et al (2009) Fig. 3d: Ubiquitin addtion

Figure 3d: Kinetics of ubiquitin addition and elongation2(Click for a larger version)


  1. Ubiquitin being a small, abundant protein[]
  2. Pierce, N., Kleiger, G., Shan, S., & Deshaies, R. (2009). Detection of sequential polyubiquitylation on a millisecond timescale Nature, 462 (7273), 615-619 DOI: 10.1038/nature08595[][][]