Abstract
Many proteins exist and function as homodimers. Understanding the detailed mechanism driving the homodimerization is important and will impact future studies targeting the "undruggable" oncogenic protein dimers. In this study, we used 14-3-3σ as a model homodimeric protein and performed a systematic investigation of the potential roles of amino acid residues in the interface for homodimerization. Unlike other members of the conserved 14-3-3 protein family, 14-3-3σ prefers to form a homodimer with two subareas in the dimeric interface that has 180° symmetry. We found that both subareas of the dimeric interface are required to maintain full dimerization activity. Although the interfacial hydrophobic core residues Leu(12) and Tyr(84) play important roles in 14-3-3σ dimerization, the non-core residue Phe(25) appears to be more important in controlling 14-3-3σ dimerization activity. Interestingly, a similar non-core residue (Val(81)) is less important than Phe(25) in contributing to 14-3-3σ dimerization. Furthermore, dissociating dimeric 14-3-3σ into monomers by mutating the Leu(12), Phe(25), or Tyr(84) dimerization residue individually diminished the function of 14-3-3σ in resisting drug-induced apoptosis and in arresting cells at G2/M phase in response to DNA-damaging treatment. Thus, dimerization appears to be required for the function of 14-3-3σ.
Highlights
Many proteins exist and function as homodimers
In a recent study analyzing the mechanisms of 14-3-3 dimerization using molecular dynamics (MD)4 simulations and site-directed mutagenesis [19], we found that Phe25 is critical in packing and stabilizing hydrophobic core residues that are important for dimerization by organizing cooperativity of core and other residues for favorable hydrophobic and electrostatic interactions
Analysis of the Dimeric Interface of 14-3-3—To investigate the determinant residues of 14-3-3 dimerization, we first examined the dimeric interface of 14-3-3 (Protein Data Bank codes 1YZ5 and 1YWT)
Summary
Many proteins exist and function as homodimers. Understanding the detailed mechanism driving the homodimerization is important and will impact future studies targeting the “undruggable” oncogenic protein dimers. In a recent study analyzing the mechanisms of 14-3-3 dimerization using molecular dynamics (MD) simulations and site-directed mutagenesis [19], we found that Phe is critical in packing and stabilizing hydrophobic core residues that are important for dimerization by organizing cooperativity of core and other residues for favorable hydrophobic and electrostatic interactions. This organizing activity of Phe for 14-3-3 homodimerization is provided by its unique physical location, rigidity, size, and hydrophobicity. We performed a systematic study of the amino acid residues in the hydrophobic core of the 14-3-3 dimeric interface along with residues not located in the hydrophobic core for their potential role in 14-3-3 dimerization and inves-
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