The mitochondrial respiratory Complex II or succinate:ubiquinone oxidoreductase (SQR) is an integral membrane protein complex in both the tricarboxylic acid cycle and aerobic respiration. Here we report the first crystal structure of Complex II from porcine heart at 2.4 A resolution and its complex structure with inhibitors 3-nitropropionate and 2-thenoyltrifluoroacetone (TTFA) at 3.5 A resolution. Complex II is comprised of two hydrophilic proteins, flavoprotein (Fp) and iron-sulfur protein (Ip), and two transmembrane proteins (CybL and CybS), as well as prosthetic groups required for electron transfer from succinate to ubiquinone. The structure correlates the protein environments around prosthetic groups with their unique midpoint redox potentials. Two ubiquinone binding sites are discussed and elucidated by TTFA binding. The Complex II structure provides a bona fide model for study of the mitochondrial respiratory system and human mitochondrial diseases related to mutations in this complex.

Coronavirus replication and transcription machinery involves multiple virus-encoded nonstructural proteins (nsp). We report the crystal structure of the hexadecameric nsp7-nsp8 supercomplex from the severe acute respiratory syndrome coronavirus at 2.4-angstroms resolution. nsp8 has a novel 'golf-club' fold with two conformations. The supercomplex is a unique hollow, cylinder-like structure assembled from eight copies of nsp8 and held tightly together by eight copies of nsp7. With an internal diameter of approximately 30 angstroms, the central channel has dimensions and positive electrostatic properties favorable for nucleic acid binding, implying that its role is to confer processivity on RNA-dependent RNA polymerase.

The unique coronavirus transcription/replication machinery comprised of multiple virus-encoded nonstructural proteins (nsp) plays a vital role during initial and intermediate phases of the viral life cycle. The crystal structure of mouse hepatitis virus strain A59 (MHV-A59) nsp15 is reported at 2.15-Å resolution. nsp15 is an XendoU endoribonuclease and is the first one from this family to have its structure unveiled. The MHV-A59 nsp15 monomer structure has a novel protein fold. Two nsp15 trimers form a back-to-back hexamer that is believed to be the functional unit. The structure reveals the catalytic site including the highly conserved residues His262, His277, and Lys317, which is supported by mutagenesis analysis. Gel filtration and enzyme activity assays confirmed that the hexamer is the active form for nsp15 and demonstrate the specificity of nsp15 for uridylate. The high sequence conservation of nsp15 in coronaviruses, including that of severe acute respiratory syndrome, suggests that this protein may provide a new target for the design of antiviral therapeutics.Mouse hepatitis virus (MHV) belongs to group II of the genus Coronavirus (CoV), together with bovine coronavirus, human coronavirus strain OC43, and the recently identified severe acute respiratory syndrome coronavirus (SARS-CoV) (2). The two strains of MHV that have been extensively studied to date are A59 (MHV-A59) and JHM (MHV-JHM or MHV-4). These coronavirus strains cause a variety of diseases in susceptible mice, such as enteritis, hepatitis, and panencephalitis, with acute and chronic demyelination that is histologically similar to multiple sclerosis in humans (13,14). Much of our knowledge concerning the replication mechanism of coronaviruses has been acquired from the use of MHV as a model for pathogenesis, docking and entry, receptor usage, transcription, replication, polymerase function, assembly, and release (31). MHV-A59 in particular is used extensively as a model to study the role of the immune system in virus-induced ventral nervous system demyelination. To date, the pathology of demyelination is still not clear; thus, further insights into the replication and transcription of the model are required to better use this model for further study of demyelination, replication, and transcription of other coronaviruses.MHV-A59 is a positive-strand RNA coronavirus with a genome of 31 kb in length, of which about 22 kb is encompassed by the replicase gene containing two large overlapping open reading frames (ORF), termed ORF1a and ORF1b. The replicase gene expresses two large polyproteins, pp1a (495 kDa) and pp1ab (803 kDa), where expression of pp1ab involves a Ϫ1 ribosomal frameshift just upstream of the ORF1a translation termination codon (31). Neither pp1a nor pp1ab is detected intact in MHV-infected cells, since they are cotranslationally and posttranslationally processed by three proteases (two papain-like proteases, PLP1 and PLP2, and a main protease, M pro ) into at least 14 mature nonstructural proteins (1,23,29). These nonstructural p...