Rethinking Leucine Zipper : ribonuclease activity and structural dynamics of a ubiquitous oligomerization motif

This dissertation focuses on structural, dynamic and catalytic properties of a Leucine
Zipper (LZ) motif – a family of protein oligomerization domains which belong to the
structural class of coiled coil proteins. LZ possess unique stability owing to high abundance of
leucine residues in the key positions of the oligomerization interface. This allows increased
combinatorial flexibility for the sidechains in coiled coil positions defining oligomerization
specificity, thus making LZ an ideal protein-protein interaction determinant. This potential is
reflected in the omnipresence of LZ within protein signalling pathways. Summarized in the
Chapter I, we review the structure, interaction specificity, folding characteristics and functional
diversity of LZ motifs, revealing the molecular mechanisms underlying LZ-enabled protein
signaling.
Beyond the widely acknowledged role of a protein oligomerization motif, recently it
was shown that LZ motifs from bZIP factors GCN4 and cJun are capable of catalyzing
degradation of RNA. Moreover catalytic RNase activity is conserved within full-length bZIP
factors. This discovery was made in the laboratory of Prof. Bernd Gutte (University of Zurich)
and served as a basis for the structural studies of LZ presented in this thesis. The manuscript
presented as the Chapter II summarizes the results of the initial LZ RNase studies, performed
in collaboration with Christine Deillon and Stefan Hoffman.
Our first structural trials on LZ-GCN4 employing solution NMR led to the discovery of
the x-form – a novel monomeric folding intermediate of LZ that exists in equilibrium with the
classical coiled coil state. Although marginally populated at experimental in vitro conditions,
x-form might represent a considerable fraction of the LZ structural ensemble in vivo, providing
a transient interface for specific recombination of interaction partners within bZIP networks.
Results of these studies are presented as Chapter III of this thesis. Finally, our structural NMR studies of LZ–RNA interactions have shown that the
substrate interacts with the coiled coil (dimeric) conformation, while the x-form is incapable of
binding RNA molecules. This is supported by the fact that the catalytic site is formed at the
interface of two LZ chains, and therefore is only available upon assembly of the coiled coil
dimer. Experimental data show that LZ from GCN4 and cJun differ in the topology and
catalytic properties of the active site, which points to the ability of LZ to provide a general
scaffold for assembly of catalytic sites with different properties. These results are presented in
the Chapter IV of this thesis.