Residue conservation in viral capsid assembly

To evaluate the evolutionary constraints placed on viral proteins by the structure and assembly of the capsid, we calculate Shannon entropies in the aligned sequences of 45 polypeptide chains in 32 icosahedral viruses, and relate these entropies to the residue location in the three-dimensional structure of the capsids. Three categories of residues have entropies lower than the chain average implying that they are better conserved than average: residues that are buried within a subunit (the protein core), residues that contain atoms buried at an interface between subunits (the interface core), and residues that contribute to several such interfaces. The interface core is also conserved in homomeric proteins and in transient protein-protein complexes, which have only one interface whereas capsids have many. In capsids, the subunit interfaces implicate most of the polypeptide chain: on average, 66% of the capsid residues are at an interface, 34% at more than one, and 47% at the interface core. Nevertheless, we observe that the degree of residue conservation can vary widely between interfaces within a capsid and between regions within an interface. The interfaces and regions of interfaces that show a low sequence variability are likely to play major roles in the self-assembly of the capsid, with implications on its mechanism that we discuss taking adeno-associated virus as an example.     

Urease inhibitor from Datisca cannabina linn

A new flavonoid datisdirin (1), along with eight known compounds tectochrysine, cearoin, sideroxyline, ursolic acid, corosolic acid, arjunolic acid, erythrodiol and oleanolic acid, were isolated from the ethyl acetate fraction of D. cannabina Linn. The structure of compound 1 was deduced on the basis of its spectral data. Datisdirin showed activity against the ureases enzyme.     

Hepatitis C virus core protein upregulates serine phosphorylation of insulin receptor substrate-1 and impairs the downstream akt/protein kinase B signaling pathway for insulin resistance

Chronic hepatitis C virus (HCV) infection has a significantly increased prevalence of type 2 diabetes mellitus (T2DM). Insulin resistance is a critical component of T2DM pathogenesis. Several mechanisms are likely to be involved in the pathogenesis of HCV-related insulin resistance. Since we and others have previously observed that HCV core protein activates c-Jun N-terminal kinase (JNK) and mitogen-activated protein kinase, we examined the contribution of these pathways to insulin resistance in hepatocytes. Our experimental findings suggest that HCV core protein alone or in the presence of other viral proteins increases Ser(312) phosphorylation of the insulin receptor substrate-1 (IRS-1). Hepatocytes infected with cell culture-grown HCV genotype 1a or 2a displayed a significant increase in the Ser(473) phosphorylation status of the Ser/Thr kinase protein kinase B (Akt/PKB), while Thr(308) phosphorylation was not significantly altered. HCV core protein-mediated Ser(312) phosphorylation of IRS-1 was inhibited by JNK (SP600125) and phosphatidylinositol-3 kinase (LY294002) inhibitors. A functional assay also suggested that hepatocytes expressing HCV core protein alone or infected with cell culture-grown HCV exhibited a suppression of 2-deoxy-d-[(3)H]glucose uptake. Inhibition of the JNK signaling pathway significantly restored glucose uptake despite HCV core expression in hepatocytes. Taken together, our results demonstrated that HCV core protein increases IRS-1 phosphorylation at Ser(312) which may contribute in part to the mechanism of insulin resistance.     

Structural evidence that the 32-kilodalton lipoprotein (Tp32) of Treponema pallidum is an L-methionine-binding protein

A structure-to-function approach was undertaken to gain insights into the potential function of the 32-kDa membrane lipoprotein (Tp32) of Treponema pallidum, the syphilis bacterium. The crystal structure of rTp32 (determined at a resolution of 1.85 A) shows that the organization of rTp32 is similar to other periplasmic ligand-binding proteins (PLBPs), in that it consists of two alpha/beta domains, linked by two crossovers, with a binding pocket between them. In the pocket, a molecule of L-methionine was detected in the electron density map. Residues from both domains interact with the ligand. One of the crossover regions is comprised of a 3(10)-helix, a feature not typical in other ligand-binding proteins. Sequence comparison shows strong similarity to other hypothetical methionine-binding proteins. Together, the data support the notion that rTp32 is a component of a periplasmic methionine uptake transporter system in T. pallidum.     

A series of related nucleotide analogues that aids optimization of fluorescence signals in probing the mechanism of P-loop ATPases, such as actomyosin

We have synthesized a set of ATP and ADP analogues that have a fluorophore linked to the nucleotide via the 3'-position of the ribose moiety. Combinations of three different coumarins are each attached via different length linkers. A linker based on propylenediamine increases the separation between the nucleotide and fluorophore relative to that of the previously reported ethylenediamine-linked coumarin nucleotides [Webb, M. R., and Corrie, J. E. T. (2001) Biophys. J. 81, 1562-1569]. A synthesis of 3'-amino-3'-deoxyATP is described using a combination of chemical and enzymatic procedures, mostly from published methods for synthesis of this compound but with some modifications that improved the convenience of the experimental procedures. This compound is used as a basis of a series of analogues with effectively a zero-length linker. Fluorescence properties of all these analogues are described, together with the kinetics of their interaction with rabbit skeletal myosin subfragment 1 in the presence and absence of actin. One particular analogue, deac-aminoATP [3'-(7-diethylaminocoumarin-3-carbonylamino)-3'-deoxyadenosine 5'-triphosphate], shows a 17-fold enhancement of fluorescence upon binding to this (skeletal) myosin II. As the diphosphate, it exhibits a large signal change upon dissociation from the actomyosin, with kinetics similar to those of natural ADP. The ability of this set of analogues to produce large signals indicated potential uses when scarce proteins are studied in small amounts.     

Anticoagulation for non-valvular atrial aibrillation - towards a new beginning with ximelagatran

OBJECTIVES: Ximelagatran is a novel oral direct thrombin inhibitor. It has favorable pharmacodynamic properties, with a broad therapeutic range without the need for anticoagulation monitoring. We aimed to discover whether ximelagatran offers a genuine future replacement to warfarin for patients in persistent atrial fibrillation (AF). MATERIALS AND METHODS: We provide an evidence-based review of the relative merits and disadvantages of warfarin and aspirin. We subsequently present an overview of the evidence for the utility of ximelagatran in the treatment of AF. RESULTS: Adjusted dose warfarin is recommended over aspirin for patients in AF at high risk of future stroke. Some of this benefit is partially offset by the higher bleeding risks associated with warfarin therapy. The SPORTIF III and V studies have shown that ximelagatran is not inferior to warfarin in the prevention of all strokes in patients with AF (both persistent and paroxysmal). This benefit was partially offset by the finding of a significant elevation of liver transaminases (>3 x normal) in 6% of patients. CONCLUSIONS: Current data would suggest that ximelagatran might represent a future alternative to warfarin. The lack of need for anticoagulant monitoring has been partially offset by a need for regular monitoring of liver function. Further data from randomized clinical trials is clearly needed.     

Design, synthesis, and evaluation of dioxane-based antiviral agents targeted against the Sindbis virus capsid protein

Dioxane-based antiviral agents targeted to the hydrophobic binding pocket of Sindbis virus capsid protein were designed by computer graphics molecular modeling and synthesized. Virus production using SIN-IRES-Luc and capsid assembly were monitored to evaluate antiviral activity. A compound with a three-carbon linker chain connecting two dioxane moieties inhibited virus production by 50% at a concentration of 40 microM, while (R)-hydroxymethyldioxane inhibited virus production by 50% at a concentration of 1 microM. Both compounds were not cytotoxic in uninfected BHK cells at concentrations of 1mM.