A subset of calcium‐binding S100 proteins show preferential heterodimerization

Abstract

The assembly of proteins into dimers and oligomers is a necessary step for the proper function of transcription factors, muscle proteins, and proteases. In uncontrolled states, oligomerization can also contribute to illnesses such as Alzheimer's disease. The S100 protein family is a group of dimeric proteins that have important roles in enzyme regulation, cell membrane repair, and cell growth. Most S100 proteins have been examined in their homodimeric state, yet some of these important proteins are found in similar tissues implying that heterodimeric molecules can also be formed from the combination of two different S100 members. In this work, we have established co‐expression methods in order to identify and quantify the distribution of homo‐ and heterodimers for four specific pairs of S100 proteins in their calcium‐free states. The split GFP trap methodology was used in combination with other GFP variants to simultaneously quantify homo‐ and heterodimeric S100 proteins in vitro and in living cells. For the specific S100 proteins examined, NMR, mass spectrometry, and GFP trap experiments consistently show that S100A1:S100B, S100A1:S100P, and S100A11:S100B heterodimers are the predominant species formed compared to their corresponding homodimers. We expect the tools developed here will help establish the roles of S100 heterodimeric proteins and identify how heterodimerization might alter the specificity for S100 protein action in cells.

The Zn2+ and Ca2+‐binding S100B and S100A1 proteins: beyond the myths

“…Considering that the affinities of S100B and S100A1 to form homo‐dimers or hetero‐dimer are similar (Deloulme et al ., ; Spratt et al ., ) and that S100A1 behaves as a dominant negative for the binding of Zn 2+ to S100B in the heterodimeric complex (see Section II.3), it becomes important to know if the balance in concentrations of S100B and S100A1 in cells is related to the control of heterodimer formation. Because the functions of S100 proteins are thought to rely on their interactions with target proteins (Santamaria‐Kisiel et al ., ; Zimmer & Weber, ), one can anticipate that S100B and S100A1 homodimers and S100A1/S100B heterodimers may also interact with different intracellular target proteins.…”

“…The S100 proteins now form the largest group of Ca 2+ ‐binding proteins found exclusively in vertebrates. A consistent feature of the S100 proteins is their association as homodimers and in some cases as heterodimers (Spratt et al ., ). The human genome encodes 21 S100 proteins, including solitary genes for S100B on chromosome 21, S100G on the X chromosome, S100P on chromosome 4, and S100Z on chromosome 5.…”

“…A key issue is the identification of target proteins specific to the individual S100 protein isoforms. The organization of the quaternary structures of the S100 proteins (Santamaria‐Kisiel et al ., ; Spratt et al ., ) and the role of Zn 2+ (Moroz, Wilson & Bronstein, ) are also important factors in the biology of these proteins that need to be clarified. Finally, it is clear that there are common functional properties for different S100 proteins that require further characterization.…”

High throughput competitive fluorescence polarization assay reveals functional redundancy in the S100 protein family

“…As a third scenario, it is still possible that there is functional redundancy within the orphan group, but our knowledge about S100 interaction partners is more limited in this group compared to the promiscuous group. Moreover, the present study covered only S100 homodimers (and the S100G monomer), although some S100 proteins can form heterodimers [40]. As an example, the S100A8/A9 (both coming from the orphan group) can form a functional heterodimer with known interaction partners [41].…”

“…Peptide synthesis. The CapZ (265-276), NCX1 (254-265), SIP (188-202), TRPM4 (129-147) and MDM4(25)(26)(27)(28)(29)(30)(31)(32)(33)(34)(35)(36)(37)(38)(39)(40)(41)(42)(43) peptides were chemically synthesized using solid phase peptide synthesis (PS3 peptide synthesizer, Protein Technologies) with Fmoc/tBu strategy in the case of (5(6)-carboxyfluorescein) labeled and unlabeled version. Peptides were purified by RP-HPLC using a Jupiter 300 Å C 18 column (Phenomenex).…”