Extended Data Fig. 9: Positive selection in Pox1p, Tdh1/2/3p and the ETC.
From: The role of metabolism in shaping enzyme structures over 400 million years
a) Multiple sequence alignment of selected regions of Pox1p orthologs. Asterix indicates residues under positive selection in the branch highlighted with the black box on the left. Other highlighted residues interact with positively selected residues in the structure. The dark square indicates the branch containing S. cerevisiae and W. anomalous orthologs for which residues under positive selection are highlighted in (b). b) Structure of the fatty-acyl coA oxidase homodimer interface obtained by using the crystal structure of the Y. lipolytica AOx protein, ylACO1 (right), (PDB:5Y9D). This crystal structure was also used as a reference to superimpose S. cerevisiae Pox1p structure from Alphafold2 (left). The positively selected amino acids Lys580 and Gln653 as well as the coordinating amino acids Tyr385 and Asn631 are highlighted. The red and the orange surfaces denote different protein chains of the homodimeric complex. The inset table denotes the residues present for various orthologs. Grey shading in the table denotes residues and branches that were under positive selection. c) Multiple sequence alignment of GAPDH enzymes in the region surrounding the K101 residue positively selected in the branch indicated by the dark square. d) Left: Tetrameric structure of GAPDH 3 from S. cerevisiae (PDB:3PYM) with positively selected residues highlighted in white. Right: Zoom into the binding site (predicted structure) with cofactor NAD and ligand GAP extracted from PDB:5JYA. The positively selected Lys101 as well as the coordinating Glu102 and the catalytically important Cys150 are highlighted. e) Multiple sequence alignment of Qcr2 highlighting residues detected as under positive selection in designated branches. f) Connectivity network of the cytochrome-c-oxidase/cytochrome bc1 complex (PDB: 6HU9). The node size indicates the length of the protein chain. Square nodes denote mitochondrially-encoded genes. The nodes are coloured with respect to the conservation ratio, where grey denotes missing orthogroups in our data selection. The edges indicate physical interactions between the proteins and edge width indicates the number of contacts between the surfaces of the two protein chains. The edges are coloured according to the number of positively selected amino acids from either protein chain on the surface between the two protein chains. g) Crystal structure of the cytochrome-c-oxidase/cytochrome bc1 complex. The structure consists of a homodimer made up of two ubiquinol cytochrome c reductase (complex III) complexes surrounded by two cytochrome c oxidase (Complex IV) complexes. Protein chains are coloured with respect to their conservation rate (left part) or chain ID (right part). Red dots denote positions where positive selection was detected in at least one branch. In addition, four interfaces are shown in detail, highlighting positively selected residues between the two surfaces. The respective positions in the complex are denoted in coloured boxes. Multiple sequence alignment illustrations created with Jalview.
