HCV IRES manipulates the ribosome to promote the switch from translation initiation to elongation.

The internal ribosome entry site (IRES) of the hepatitis C virus (HCV) drives noncanonical initiation of protein synthesis necessary for viral replication. Functional studies of the HCV IRES have focused on 80S ribosome formation but have not explored its role after the 80S ribosome is poised at the start codon. Here, we report that mutations of an IRES domain that docks in the 40S subunit's decoding groove cause only a local perturbation in IRES structure and result in conformational changes in the IRES-rabbit 40S subunit complex. Functionally, the mutations decrease IRES activity by inhibiting the first ribosomal translocation event, and modeling results suggest that this effect occurs through an interaction with a single ribosomal protein. The ability of the HCV IRES to manipulate the ribosome provides insight into how the ribosome's structure and function can be altered by bound RNAs, including those derived from cellular invaders

Health-related quality of life after myocardial infarction is associated with level of left ventricular ejection fraction

<p>Abstract</p> <p>Background</p> <p>The objective was to explore the relationship between left ventricular ejection fraction (LVEF) assessed during hospitalization for acute myocardial infarction (MI) and later health-related quality of life (HRQoL).</p> <p>Methods</p> <p>We used multivariable linear regression to assess the relationship between LVEF and HRQoL in 256 MI patients who responded to the Kansas City Cardiomyopathy Questionnaire (KCCQ), the EQ-5D Index, and the EuroQol Visual Analogue Scale (EQ-VAS) 2.5 years after the index MI.</p> <p>Results</p> <p>167 patients had normal LVEF (>50%), 56 intermediate (40%–50%), and 33 reduced (<40%). The mean (SD) KCCQ clinical summary scores were 85 (18), 75 (22), and 68 (21) (<it>p </it><0.001) in the three groups, respectively. The corresponding EQ-5D Index scores were 0.83 (0.18), 0.72 (0.27), and 0.76 (0.14) (<it>p </it>= 0.005) and EQ-VAS scores were 72 (18), 65 (21), and 57 (20) (<it>p </it>= 0.001). In multivariable linear regression analysis age ≥ 70 years, known chronic obstructive pulmonary disease (COPD), subsequent MI, intermediate LVEF, and reduced LVEF were independent determinants for reduced KCCQ clinical summary score. Female sex, medication for angina pectoris at discharge, and intermediate LVEF were independent determinants for reduced EQ-5D Index score. Age ≥ 70 years, COPD, and reduced LVEF were associated with reduced EQ-VAS score.</p> <p>Conclusion</p> <p>LVEF measured during hospitalization for MI was a determinant for HRQoL 2.5 years later.</p

Simvastatin Sodium Salt and Fluvastatin Interact with Human Gap Junction Gamma-3 Protein

Finding pleiomorphic targets for drugs allows new indications or warnings for treatment to be identified. As test of concept, we applied a new chemical genomics approach to uncover additional targets for the widely prescribed lipid-lowering pro-drug simvastatin. We used mRNA extracted from internal mammary artery from patients undergoing coronary artery surgery to prepare a viral cardiovascular protein library, using T7 bacteriophage. We then studied interactions of clones of the bacteriophage, each expressing a different cardiovascular polypeptide, with surface-bound simvastatin in 96-well plates. To maximise likelihood of identifying meaningful interactions between simvastatin and vascular peptides, we used a validated photo-immobilisation method to apply a series of different chemical linkers to bind simvastatin so as to present multiple orientations of its constituent components to potential targets. Three rounds of biopanning identified consistent interaction with the clone expressing part of the gene GJC3, which maps to Homo sapiens chromosome 7, and codes for gap junction gamma-3 protein, also known as connexin 30.2/31.3 (mouse connexin Cx29). Further analysis indicated the binding site to be for the N-terminal domain putatively ‘regulating’ connexin hemichannel and gap junction pores. Using immunohistochemistry we found connexin 30.2/31.3 to be present in samples of artery similar to those used to prepare the bacteriophage library. Surface plasmon resonance revealed that a 25 amino acid synthetic peptide representing the discovered N-terminus did not interact with simvastatin lactone, but did bind to the hydrolysed HMG CoA inhibitor, simvastatin acid. This interaction was also seen for fluvastatin. The gap junction blockers carbenoxolone and flufenamic acid also interacted with the same peptide providing insight into potential site of binding. These findings raise key questions about the functional significance of GJC3 transcripts in the vasculature and other tissues, and this connexin’s role in therapeutic and adverse effects of statins in a range of disease states

Expression, Purification, Structural and Functional Analysis of SycB: A Type Three Secretion Chaperone From Yersinia Enterocolitica

In Yersinia enterocolitica biovar 1B, a genome encoded TTSS designated as Ysa-Ysp system is used for virulence. SycB is an annotated chaperone to this system. SycB is soluble in presence of translocator YspC. SycB and its truncated form (DSycB(1–114)) exist as dimers. YspC forms a 1:1 complex with SycB. Homology model of SycB shows a flexible N-terminal may be required for solubility and dimerization; and concave core formed by antiparallel helices of TPRs. Far UV CD spectra confirm that SycB is predominantly alpha helical. Near UV CD spectra show that SycB has tertiary structure at pH 7.2 (native folded protein), which disappears at pH 5 (molten globule) and SycB releases YspC at pH 5. SycB has a cooperative melting behavior. At pH 7.2, SycB shows solvent accessible hydrophobic patches. Concave core in the model exhibits ANS binding within FRET distance of tyrosines in the TPR, allowing a range of interaction of SycB with its ligand

Cryo-EM structure of the αvβ8 integrin reveals a mechanism for stabilizing integrin extension

Integrins are conformationally flexible cell surface receptors that survey the extracellular environment for their cognate ligands. Interactions with ligands are thought to be linked to global structural rearrangements involving transitions between bent, extended-closed and extended-open forms. Thus far, structural details are lacking for integrins in the extended conformations due to extensive flexibility between the headpiece and legs in this conformation. Here we present single-particle electron cryomicroscopy structures of human αvβ8 integrin in the extended-closed conformation, which has been considered to be a low-affinity intermediate. Our structures show the headpiece rotating about a flexible αv knee, suggesting a ligand surveillance mechanism for integrins in their extended-closed form. Our model predicts that the extended conformation is mainly stabilized by an interface formed between flexible loops in the upper and lower domains of the αv leg. Confirming these findings with the αvβ3 integrin suggests that our model of stabilizing the extended-closed conformation is generalizable to other integrins