the hydrophobic environment of the quinone-binding pocket Qp. In contrast, ubiquinone binding site QD, which lies closer to inter-membrane space, is composed of SdhD only and has lower affinity to ubiquinone. Succinate binding site SdhA provides the binding site for the oxidation of succinate. The side chains Thr254, His354, and Arg399 of subunit A stabilize the molecule while FAD oxidizes and carries the electrons to the first of the iron-sulfur clusters, [2Fe-2S]. This can be seen in image 5. Redox centers The succinate-binding site and ubiquinone-binding site are connected by a chain of redox centers including FAD and the iron-sulfur clusters. This chain extends over 40 Å through the enzyme monomer. All edge-to-edge distances between the centers are less than the suggested 14 Å limit for physiological electron transfer. This electron transfer is demonstrated in image 8. Assembly and maturation All subunits of human mitochondrial SDH are nuclear encoded. After translation, SDHA subunit is translocated as apoprotein into the mitochondrial matrix. Subsequently, one of the first steps is covalent attachment of the FAD cofactor (covalent flavinylation). This process is enhanced by succinate dehydrogenase assembly factor 2 (SDHAF2; also called Sdh5 in yeast and SdhE in bacteria) and by some of the Krebs cycle intermediates. Fumarate most strongly stimulates covalent flavinylation of SDHA. Through studies of the bacterial system, the mechanism of FAD attachment has been shown to involve a quinone:methide intermediate. In mitochondrial, but not bacterial, assembly, SDHA interacts with a second assembly factor called succinate dehydrogenase assembly factor 4 (SDHAF4; called Sdh8 in yeast) before it is inserted into the final complex. Fe-S prosthetic groups of the subunit SDHB are being preformed in the mitochondrial matrix by protein complex ISU. The complex is also thought to be capable of inserting the iron-sulphur clusters in SDHB during its maturation. The studies suggest that Fe-S cluster insertion precedes SDHA-SDHB dimer forming. Such incorporation requires reduction of cysteine residues within active site of SDHB. Both reduced cysteine residues and already incorporated Fe-S clusters are highly susceptible to ROS damage. Two more SDH assembly factors, SDHAF1 (Sdh6) and SDHAF3 (Sdh7 in yeast), seem to be involved in SDHB maturation in way of protecting the subunit or dimer SDHA-SDHB from Fe-S cluster damage caused by ROS. Assembly of the hydrophobic anchor consisting of subunits SDHC and SDHD remains unclear. Especially in case of heme b insertion and even its function. Heme b prosthetic group