The proteasome is the central enzyme of the
ATP-ubiquitin-dependent proteolytic pathway. It is
responsible for the elimination of abnormal proteins, arising
from mutations or damage ensuing from adverse external
conditions. Moreover, it has important regulatory roles in
numerous biological processes such as cell cycle, apoptosis
or the immune response. A research team at the Max Planck
Institute of Biochemistry in Martinsried/Germany, headed by
Wolfgang Baumeister, since several years one of the leading
groups in the proteasome field, has published a
review-article in Cell on February 6, 1998, reflecting
on the proteasome as one of the most fascinating
macromolecular machines of the cell. The authors report
on own data and review the current status of the topic.
Protein degradation is a necessity but also a hazard. It must
be subject to precise spatial and temporal control to prevent
that proteins not destined for degradation are attacked. A
basic strategy in controlling protein degradation is
compartmentalization, that is sequestration of proteolytic
action from the environment. Many proteases reside within
spezialized membrane-surrounded organelles, such as
lysosomes. Another mechanism of control, which offers greater
logistic flexibility, is self- or autocompartmentalization:
proteolytic subunits self-associate into barrel-shaped
complexes with inner cavities that harbour the active sites.
The advances made in recent years in understanding the
structure of the proteasome and its mechanism of action has
helped to shape the concept of self-compartmentalization, and
the proteasome became its paradigm.
The molecular species that functions in the
ubiquitin-proteasome pathway is the so-called 26S proteasome,
a large complex composed of approx. 35 different subunits. A
smaller particle, the 20S proteasome, forms the core of the
proteolytic machinery. First observed in the late sixties in
human blood cell and subsequently isolated from numerous
eukaryotic organisms, it was, about ten years ago, also found
in an archeabacterium, Thermoplasma acidophilum. The
recent discovery of 20S proteasomes in a group of bacteria
shows that it is in fact an ancestral particle common to all
domains of life.
The molecular architecture of the 20S proteasome is conserved
from archaea to eukarya, inspite of an increasing complexity
in subunit composition. Whereas the Thermoplasma
proteasome is built from only two subunit, a and b,
eukaryotic proteasomes contain 14 different subunits; based
on sequence similarity they can be classified as a-type and b-type.
The subunits are arranged in four seven-membered rings, a-type subunits forming the outer, b-type forming the central rings.
Collectively, they build up a cylinder, traversed by a
channel which widens into three large cavities that are
separated by constrictions. In the Thermoplasma
proteasome the two outermost constrictions are so narrow that
only unfolded polypeptides are admitted to the interior. This
size exclusion mechanism was visualized by electron
microscopy with a gold labeled insulin b-chain
caught in transit; the bulky gold label prevents the insulin
chain from passing the constriction.
The enzymatic mechanism of the proteasome has been a mystery
until crystal structure analysis and mutagenesis studies on
the Thermoplasma proteasome in the laboratories of
Robert Huber and Wolfgang Baumeister threw light onto it. The
N-terminal threonine was identified as the long-sought
catalytic nucleophile.
While the Thermoplasma proteasome has taken a pivotal
role in structural and functional studies, the proteasome
from the eubacterium Rhodococcus offered an excellent
means to dissect the assembly pathway of the complex. The
team of Wolfgang Baumeister isolated and characterized the
first bacterial proteasome, and managed to set up a system,
which allows to monitor its biogenesis in vitro. Rhodococcus
a- and b-subunits
spontaneously associate when brought together, and form half
proteasomes. These inactive intermediates dimerize to
preholoproteasomes, which in turn are converted to active
holoproteasomes through the autocatalytic cleavage of the b-propeptide. The propeptide not only
prevents premature activation by blocking the N-terminus, it
also serves to support the initial folding of b-subunits.
Two complexes are found to associate with the 20S proteasome.
The 11S regulator, or PA28, binds to the outer rings of the
proteasome and modulates its activity; the biological
function of PA28 is presumed to be related to the
proteasome«s role in antigen processing. Recently,
Christopher Hill and coworkers from the university of Utah,
Salt Lake City, have solved the crystal structure of the
heptameric complex, which is traversed by a funnel-shaped
channel, the wider opening oriented to the proteasome. The
second complex when associated with the 20S proteasome yields
the 26S proteasome. The so called 19S caps are flexibly
linked to one or both ends of the 20S core and consist of
about 20 proteins. The caps serve to recognize proteins to be
degraded and to prepare them for degradation. However, the
precise function of the majority of the subunits remain
enigmatic, and also the subunit topology is at present
unknown. The challenge for the years to come is to understand
the cascade of events that starts with the binding of
ubiquitylated substrate protein and ends with its
translocation into the 20S proteolytic core.
Journal
Cell