Key result
NMR spectroscopy revealed that the isolated N-domain of Cdc4p retains its native structure, whereas the C-domain is less stable, more dynamic, and structurally perturbed in isolation.
The C-domain of Cdc4p exhibits conformational plasticity, which may allow it to interact with various regulatory or contractile ring proteins during cytokinesis.
C-domain instability in Cdc4p is hypothesis-generating for cytokinesis; leaves open in vivo roles in contractile protein regulation.
Cytokinesis is the process by which one cell divides into two. Key in the cytokinetic mechanism of Schizosaccharomyces pombe is the contractile ring myosin, which consists of two heavy chains (Myo2p), two essential light chains (Cdc4p), and two regulatory light chains (Rlc1p). Cdc4p is a dumbbell-shaped EF-hand protein composed of N- and C-terminal domains separated by a flexible linker. The properties of these two domains are of particular interest because each is hypothesized to have independent functions in binding different components of the cytokinesis machinery. To help define these properties, we used NMR spectroscopy to compare the structure, stability, and dynamics of the isolated N- and C-terminal domains with one another and with native Cdc4p. On the basis of invariant chemical shifts, the N-domain retains the same structure in isolation as in the context of the full-length Cdc4p, whereas the C-domain appears markedly perturbed. This perturbation results from intramolecular binding of the residual linker sequence at the N-terminus of the C-domain in a mode similar to that used by native Cdc4p to associate with target polypeptide sequences. NMR relaxation, thermal denaturation, and amide hydrogen exchange experiments also indicate that the C-domain is less stable and more dynamic than the N-domain, both in isolation and in the full-length protein. We hypothesize that these properties reflect a conformational plasticity of the C-domain, which may allow Cdc4p to interact with several regulatory or contractile ring proteins necessary for cytokinesis.
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Escobar-Cabrera et al. (2005) studied Schizosaccharomyces pombe cytokinesis. Isolated N- and C-terminal domains of Cdc4p vs. Native full-length Cdc4p was evaluated on Structure, stability, and dynamics assessed by NMR spectroscopy. NMR spectroscopy revealed that the isolated N-domain of Cdc4p retains its native structure, whereas the C-domain is less stable, more dynamic, and structurally perturbed in isolation.
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