DOM-fold: a structure with crossing loops found in DmpA, ornithine acetyltransferase, and molybdenum cofactor-binding domain

Abstract

Understanding relationships between sequence, structure, and evolution is important for functional characterization of proteins. Here, we define a novel DOM-fold as a consensus structure of the domains in DmpA (L-aminopeptidase D-Ala-esterase/amidase), OAT (ornithine acetyltransferase), and MocoBD (molybdenum cofactor-binding domain), and discuss possible evolutionary scenarios of its origin. As shown by a comprehensive structure similarity search, DOM-fold distinguished by a two-layered beta/alpha architecture of a particular topology with unusual crossing loops is unique to those three protein families. DmpA and OAT are evolutionarily related as indicated by their sequence, structural, and functional similarities. Structural similarity between the DmpA/OAT superfamily and the MocoBD domains has not been reported before. Contrary to previous reports, we conclude that functional similarities between DmpA/OAT proteins and N-terminal nucleophile (Ntn) hydrolases are convergent and are unlikely to be inherited from a common ancestor.

Figure 1.

Structural and topological comparisons of DmpA and Ntn hydrolase. MOLSCRIPT (Kraulis 1991) and topology diagrams of DmpA (A) and archaeal proteasome β subunit (Groll et al. 2003) as an example of Ntn hydrolases (B). In DmpA diagrams, α-helices and β-strands constituting the two structural domains, DmpA_1 and DmpA_2, are colored from blue (N terminus) to red (C terminus). The remaining secondary structure elements are shown in gray. α-Helices and β-strands are labeled with upper- or lowercase letters, respectively, followed by the domain number (e.g., strand a in DmpA_1 is labeled a1 while strand a in DmpA_2 is labeled a2). Labels are in the same color as the secondary structure elements they represent. Except for the crossing loops highlighted in green and red, loop regions are shown in gray. Dotted lines indicate the autocleavage site. In Ntn hydrolase diagrams, the secondary structure elements are colored and labeled according to their spatial equivalents in DmpA. The last α-helix and β-strand are colored white since they do not have structural equivalents in DmpA. For all structures, N and C termini are labeled and the PDB code, the chain ID, and the starting and ending residue numbers are indicated. Catalytic nucleophiles (Ser250 in DmpA and Thr1 in Ntn hydrolase) are shown in ball-and-stick representation or as asterisks in MOLSCRIPT or topology diagrams, respectively. To make the topology diagrams, each structure is separated between the two β-sheets and rotated roughly 90° as indicated by the curved arrows (“open book”).

Figure 2.

Structural comparison of DmpA and MocoBD domains. (A) The two structural domains of DmpA. (B) The structure of MocoBD. (C) The four domains of MocoBD. In A and C, spatially equivalent secondary structure elements are colored and labeled correspondingly in the same way as in Figure 1A ▶. In B, the four structural domains are shown in different colors: 1N, light yellow; 1C, dark yellow; 2N, light blue; 2C, dark blue. The two lobes are colored in yellow and blue, respectively. The molybdenum cofactor in B and C is shown in bonds representation with the molybdenum atom highlighted as a black ball. In C, the rectangular bar below each structure shows the color in which that domain is represented in B. For all structures, the crossing loops are highlighted in green and red; secondary structure elements that do not belong to the consensus fold of DmpA and MocoBD domains are shown in gray; dotted lines represent disordered regions or long insertions; N and C termini are labeled and the PDB code, the chain ID, and the starting and ending residue numbers are shown. Diagrams are drawn with MOLSCRIPT (Kraulis 1991). (D) A proposed model for MocoBD to evolve from a single-domain ancestor. The single-domain ancestor may form a homodimer (or a homotetramer). Duplication, fusion, and domain swapping created one lobe in which the two duplicates exchange their first strand. The linker region between the two domains is colored in blue. The lower, mainly red domain corresponding to 1N/2N seems to be circularly permuted and inserted into the upper, mainly green domain corresponding to 1C/2C. Further duplication of this lobe resulted in the four domains in MocoBD. The linker between the two lobes is colored in cyan.

Figure 3.

Multiple sequence alignment of representative sequences in DmpA/OAT superfamily (A) and MocoBD superfamily (B). Each group of sequences corresponds to a structural domain specified to the left of the group. Different sequences within the same group were aligned by the PCMA program (Pei et al. 2003). For each sequence, the NCBI gene identification (gi) number follows the serial number; the first and last residue numbers are indicated before and after the sequence, respectively, and the full length of the protein is shown in parentheses at the end. The species name abbreviations follow the gi numbers and are colored in red, black, and green for archaea, bacteria, and eukaryota, respectively. The eight groups were aligned together according to a structural superposition of the eight domains in the three reference structures (PDB 1b65 chain A for DmpA, PDB 1vz6 chain A for OAT, and PDB 1jro chain B for MocoBD). The structural features used for selecting the registers are marked by various symbols (*, @, #, &). The first gi number in each group is underlined to indicate that it corresponds to the reference structure. Secondary structure elements (E for β-strands and H for α-helices) are labeled above each group according to the reference structure. Note that MocoBD_1N and MocoBD_1C or MocoBD_2N and MocoBD_2C as shown in Figure 2C ▶ swapped their first strands in this multiple alignment to make the sequences continuous, as indicated by the red boxes and arrows. The secondary structure diagram (cylinders for α-helices and arrows for β-strands) at the bottom of this figure is colored and labeled in the same way as in Figure 2, A and C ▶. Uncharged residues (any residue except K, R, E, D) at positions in the interface between the α-helix layer and the β-strand layer in each domain are shaded in yellow. Small residues (G, A, C, P, T, S, V, D, N) at positions containing mainly small residues are shaded in gray. Long insertions between the aligned secondary structure elements are omitted for clarity, and the numbers of omitted residues are indicated in parentheses. In A, the autocleavage site is indicated by a black arrow. The functional residues of DmpA and OAT are identified according to Bompard-Gilles et al. (2000) and Elkins et al. (2005), respectively, and are highlighted in different colors: nucleophile, black; nucleophile stabilization, cyan; and oxyanion hole, green. In B, the residues participating in molybdenum cofactor-binding are identified according to Figure 6 in Truglio et al. (2002) and are highlighted in purple. Species name abbreviations are At, Arabidopsis thaliana; Bs, Bacillus stearothermophilus; Bt, Bos taurus; Ca, Chloroflexus aurantiacus; Dg, Desulfovibrio gigas; Fl, Flavobacterium sp.; Kl, Kluyveromyces lactis NRRL Y-1140; Mg, Magnaporthe grisea 70-15; Mj, Methanocaldococcus jannaschii DSM 2661; Ml, Mesorhizobium loti MAFF303099; Oa, Ochrobactrum anthropi; Oc, Oligotropha carboxidovorans; Pa, Pyrococcus abyssi GE5; Pp, Pseudomonas putida; Rc, Rhodobacter capsulatus; Sc, Saccharomyces cerevisiae; Ss, Sulfolobus solfataricus P2; St, Streptomyces clavuligerus; Su, Sulfolobus tokodaii str. 7; Tn, Thermotoga neapolitana; Tp, Treponema pallidum subsp. pallidum str. Nichols; Tt, Thermoanaerobacter tengcongensis.