Heme-Copper Oxygen Reductase

Heme-copper Oxygen reductases (HCOs) are the main enzymes responsible for the reduction of oxygen to water in respiratory chains. These membrane-bound enzymes are present in the three life domains, Bacteria, Archaea and Eukarya, catalyzing the last reaction of aerobic respiratory chains. HCOs contribute to energy conservation by two processes: i) charge separation, since protons and electrons needed for the chemical reaction come from opposite sides of the membrane, and ii) proton translocation, as part of the energy released during the O2 reduction is used to promote thermodynamically unfavorable proton translocation across the membrane.

Sequence alignments, site directed mutagenesis and X-ray crystallographic structural models led to the identification of intra-protein proton conducting channels in the mitochondrial and mitochondrial-like enzymes. These channels were named D and K due to specific amino acid residues considered to play an important role in proton translocation.

We have observed that those specific residues were not conserved in all HCOs and we have indicated possible alternatives in different enzymes. Based on the fingerprint of the proton conducting channels we have proposed a classification scheme in which HCOs were divided into three types: A (further divided into A1 and A2), B and C Type enzymes. This classification was recently strengthened by a comprehensive bioinformatics study.

The A Type enzymes are the highest number of proteins known so far and are the best studied HCOs. This group includes the mitochondrial oxygen reductase and is characterized by the presence of two proton conducting channels (the D- and K-channel) and by the location of the catalytic tyrosine in helix VI (Tyr-I). In B Type enzymes only one identified proton conducting channel has been identified, the alternative K-channel, and in the majority of those enzymes the catalytic tyrosine is also present in helix VI (Tyr-I). C Type HCOs also contain only one channel, another alternative K-channel, characteristic of this group and different from that of B Type enzymes. The catalytic tyrosine residue is located in helix VII (Tyr-II).

In collaboration with the Computational Genomics Laboratory from Instituto Gulbenkian de Ciência a tool able to classify any user supplied sequence was created. This involved the built up of a pipeline where new incoming sequences were classified according to their best similarity hit against a previously manually curated dataset. The method of this automated classification relies on the similarities between amino acid sequences obtained by local alignments (blast) against the dataset presented in this work. This algorithm is available as a web-tool at http://www.evocell.org/hco/.