1. Structure and Function of an Unusual Family of Protein Phosphatases
Sang-Youn Park, Xingjuan Chao, Gabriela Gonzalez-Bonet, Bryan D. Beel, Alexandrine M. Bilwes, Brian R. Crane 
Molecular Cell 
Volume 16, Issue 4, Pages (November 2004)
DOI: /j.molcel

2. Figure 1
The CheC Family
(A) Domain organization for the family of protein aspartate-phosphatases and related proteins in three different bacteria, T. maritima, B. subtilis, and E. coli. Purple segments represent the CheC homology region; green segments represent the FliN homology region (PDB code: 1O6A). FliY/N and FliM contain an N-terminal peptide that binds CheY-P (black). CheC, CheX, and FliY/N contain dephosphorylation centers (white stars with conserved residues above), but FliM does not. Most FliY/N proteins follow the domain architecture of bsFliY/N. TmFliY is abnormally short. The structurally unrelated CheY-phosphatase CheZ found in β- and γ-proteobacteria is shown in red.
(B) Sequence alignments of T. maritima CheC, FliY, and CheX. Secondary structure elements of CheC (above) and CheX (below) are similar except for the regions (in red) that form helices in CheC (α2 and α2′) and β strands (βX and βX′) in the dimer interface of CheX. Residue conservation (boxes) clusters in the regions of α1 (α1′) and β1 (β1′). Black boxes highlight conserved residues in the active site regions, and red boxes encircle residues that are markers for the CheX family versus CheC family.
Molecular Cell  , DOI: ( /j.molcel )

3. Figure 2 The Folds of CheC and CheX
(A) Ribbon diagrams show topologies and secondary structural elements for the CheC monomer (left) and one CheX subunit (right) from two orientations. The approximate locations of important invariant residues (e.g. Glu13, Asn16, and Pro39 for CheC, left active site) are shown by stars on white circles. Pseudo-2-fold axes perpendicular to the page in upper image relate one half of each molecule to the other. The kink in β1 (β1′) wraps the center of α1 (α1′) where conserved active site residues reside. Lower images show how CheC α2 and α2′ replaced βX and βX′ that form the dimer interface in CheX.
(B) Dimer formation in CheX. Two continuous β sheets associate the CheX subunits (blue and gray). Strands βX′ from each subunit (light and dark orange) swap across the dimer interface to form main chain hydrogen bonds with β1′ of the adjacent subunit. The flagella proteins, FliY/N and FliM are predicted to have structures more analogous to CheC than CheX.
(C) Solvent-accessible surfaces for CheC (left) and CheX (right) rendered transparent to view protein topology and key residue structure within. Conserved proline residues (white bonds) on β1′ lie at the center of the CheX dimer interface (blue and gray subunits, yellow and green ribbons) adjacent to the conserved Glu and Asn residues on α1′ (white bonds). In CheC, the conserved prolines (white bonds) on β1′ form a protrusion at the side of the molecule that may mediate contact with another protein. Conserved residues in the other active site of CheC (α1-β1) also project into solvent (white bonds, lower left).
Molecular Cell  , DOI: ( /j.molcel )

4. Figure 3 Activities of T. maritima CheC, CheX, and CheD
(A) Flow of phosphate followed in the experiments shown in (B)–(C).
(B) Autophosphorylated CheA (CheA-32P) in the absence or presence of CheY, CheC, CheD, CheX, and CheC double mutants Glu13Ser + Glu112Ser (CheC dmE) or Asn16Ser + Asn115Ser (CheC dmN). Only CheY dephosphorylates CheA (lane 2). Both CheX (lane 10) and CheC (lane 6) reduce the amount of CheY-P, although CheX has much greater activity as no CheY-P remains in its presence. CheD activates CheC (lane 12) to roughly the same level as CheX (lane 10). CheC dmE and CheC dmN do not noticeably dephosphorylate CheY (lanes 7 and 8); however, CheD partially rescues the activity of dmE (lane 13), but not dmN (lane 14).
(C) Effects of CheC mutants on CheY-P in the presence of CheD. Bands corresponding to CheY-P after transfer from autophosphorylated CheA (CheA-32P). Lanes 1–7 are controls as designated; lanes 8–13 are CheA+CheY+CheD+CheC mutants: Glu13Ser, Glu112Ser, Glu13Ser/Glu112Ser, Asn16Ser, Asn115Ser, and Asn16Ser/Asn115Ser, respectively. Coomassie-stained SDS-PAGE gel showing that the concentrations of the CheC mutants are equivalent.
Molecular Cell  , DOI: ( /j.molcel )

5. Figure 4 The Dephosphorylation Centers of CheC and CheX
(A–D) Stereoimages of 2Fo − Fc electron-density maps (contoured at 1σ, green for α1 and α1′ and gray for β1 and β1′) showing regions surrounding conserved Asn16 (A) or Asn115 (B) for CheC; and surrounding conserved Asn94 for CheX in the A subunit of the dimer (C) and the B subunit of the dimer (D). Conformations of the conserved Asn and Glu residues slightly differ in each case, as does the juxtaposition of the kink in β1 (β1′) provided by the Pro (Val)-Pro motif. This variability is most dramatic in CheX where the two subunits differ by a shift in α1′ of one helical turn relative to the Pro-Pro repeat (C and D).
Molecular Cell  , DOI: ( /j.molcel )

6. Figure 5 Comparing the dephosphorylation centers of CheC and CheZ
Comparison between the putative active center residues of CheC and those of CheZ in complex with activated CheY. Superposition of CheZ Gln147 with CheC Glu13 and their bearing helices leads to a clash-free complex between CheY and CheC and aligns CheZ Asp143 with CheC Asp9. However, the catalytically essential CheC Asn16 does not reside in the interface; hence, recognition of CheY-P by CheC must be different than by CheZ.
Molecular Cell  , DOI: ( /j.molcel )

7. Figure 6 Helical Shifts in CheX and CheC
Stereoimage of the superposition of Cαs traces for the two CheX molecules (red and green) in our structure, the two molecules (gray) in the deposited coordinates of CheX (PDB code: 1SQU) and CheC (blue). The N-terminal Met (Met1) of CheX bridges interactions between α1, α1′, and the β sheet in all subunits except subunit A of our structure, where the N-terminus of α1 projects into solvent and as a result α1, α1′ shift by ∼one helical turn towards the dimer interface. In CheC, α3′ structurally mimics the CheX N-terminal Met, resulting in a third conformation for α1 and α1′. The side chains for the following conserved residues are represented: Asn115, Pro137, Pro138 (CheC) and Met1, Asn94, Pro115, Pro116 (CheX).
Molecular Cell  , DOI: ( /j.molcel )