Switch-of-function mutants based on morphology classification of Ras superfamily small GTPases.
Signaling proteins from the same family can have markedly different roles in a given cellular context. Here, we show that expression of one hundred constitutively active human small GTPases induced cell morphologies that fell into nine distinct classes. We developed an algorithm for pairs of classes that predicted amino acid positions that can be exchanged to create mutants with switched functionality. The algorithm was validated by creating switch-of-function mutants for Rac1, CDC42, H-Ras, RalA, Rap2B, and R-Ras3. Contrary to expectations, the relevant residues were mostly outside known interaction surfaces and were structurally far apart from each other. Our study shows that specificity in protein families can be explored by combining genome-wide experimental functional classification with the creation of switch-of-function mutants.
Heo
Won Do
WD
Department of Molecular Pharmacology, Stanford University School of Medicine, 269 Campus Drive, Room 3215, Stanford, CA 94305, USA.
Meyer
Tobias
T
D016475:3T3 Cells; D000465:Algorithms; D000595:Amino Acid Sequence; D000818:Animals; D048429:Cell Size; D006367:HeLa Cells; D006801:Humans; D051379:Mice; D008958:Models, Molecular; D020559:Monomeric GTP-Binding Proteins / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism*; D009154:Mutation / Q000235:genetics*; D011487:Protein Conformation; D017386:Sequence Homology, Amino Acid; D013329:Structure-Activity Relationship
D020559:Monomeric GTP-Binding Proteins