Polymerization of the SAM domain of MAPKKK Ste11 from the budding yeast: implications for efficient signaling through the MAPK cascades.
The sterile alpha-motif (SAM) is a protein module approximately 70 residues long and mainly involved in the protein-protein interactions of cell signaling and transcriptional repression. The SAM domain of the yeast MAPKKK Ste11 has a well-folded dimeric structure in solution. Interestingly, the well-folded dimer of the Ste11 SAM undergoes a time-dependent self-assembly upon lowering of the pH, leading to the formation of high molecular weight oligomers. The oligomeric structures rapidly disassemble to the well-folded dimer upon reversal of the pH to close to neutral conditions. Circular dichroism (CD) and atomic force microscopy (AFM) experiments demonstrate that the oligomeric structure formed at pH 5.0 appears to be highly helical and has architecture akin to proto-fibrils. Residue-specific kinetics of pH-triggered oligomerization obtained from real-time 15N-1H HSQC experiments indicate that the dimer-oligomer transition appears to involve all residues of the well-folded dimeric structure of the Ste11 SAM. Very interestingly, the interactions of the Ste11 and Ste50 SAM domains also lead to the formation of non-homogeneous hetero-complexes with significant populations of high molecular weight aggregates. AFM imaging shows that the Ste11-Ste50 hetero-polymeric aggregates assume the shapes of circular nano-particles with dimensions of 50-60 nano-meters (nm), in contrast to the proto-fibrils formed by the Ste11 SAM domain alone. Such intrinsic propensity for dimer to oligomer transition of the Ste50-binding SAM domain of Ste11 may endow the MAPKKK Ste11 with unique functional properties required for efficient and high fidelity signal transduction in the budding yeast.
Bhattacharjya
Surajit
S
Biomolecular NMR and Protein Research, Biotechnology Research Institute, National Research Council of Canada, 6100 Royalmount Avenue, Montreal, Quebec H4P 2R2, Canada.
Xu
Ping
P
Chakrapani
Mukundan
M
Johnston
Linda
L
Ni
Feng
F
D002942:Circular Dichroism; D019281:Dimerization; D006863:Hydrogen-Ion Concentration; D020930:MAP Kinase Kinase Kinases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D018625:Microscopy, Atomic Force; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism*; D025941:Protein Interaction Mapping; D017434:Protein Structure, Tertiary; D029701:Saccharomyces cerevisiae Proteins / Q000378:metabolism; D004718:Saccharomycetales / Q000201:enzymology; D015398:Signal Transduction / Q000502:physiology
C079506:STE50 protein, S cerevisiae; D029701:Saccharomyces cerevisiae Proteins; D020928:Mitogen-Activated Protein Kinases; D020930:MAP Kinase Kinase Kinases; C067018:Ste11 protein, S cerevisiae