Conformational changes in tubulin upon binding cryptophycin-52 reveal its mechanism of action

Cryptophycin-52 (Cp-52) is potentially the most potent anticancer drug known, with IC₅₀ values in the low picomolar range, but its binding site on tubulin and mechanism of action are unknown. Here, we have determined the binding site of Cp-52, and its parent compound, cryptophycin-1, on HeLa tubulin, to a resolution of 3.3 Å and 3.4 Å, respectively, by cryo-EM and characterized this binding further by molecular dynamics simulations. The binding site was determined to be located at the tubulin interdimer interface and partially overlap that of maytansine, another cytotoxic tubulin inhibitor. Binding induces curvature both within and between tubulin dimers that is incompatible with the microtubule lattice. Conformational changes occur in both α-tubulin and β-tubulin, particularly in helices H8 and H10, with distinct differences between α and β monomers and between Cp-52-bound and cryptophycin-1-bound tubulin. From these results, we have determined: (i) the mechanism of action of inhibition of both microtubule polymerization and depolymerization, (ii) how the affinity of Cp-52 for tubulin may be enhanced, and (iii) where linkers for targeted delivery can be optimally attached to this molecule.     

Fixed-bed catalytic pyrolysis of cotton-seed cake: effects of pyrolysis temperature, natural zeolite content and sweeping gas flow rate

Catalyzed pyrolysis of cotton-seed cake was studied under different experimental conditions. Variables investigated were pyrolysis temperature, zeolite content and sweeping gas flow rate. Experiments were carried out isothermally. Liquids, gases and char were obtained as products of pyrolysis. The distributions of these products were determined for various contents (1, 5, 10, 20 wt.% of raw material) of zeolite at four different pyrolysis temperatures. The maximum liquid yield obtained was 30.84% at a pyrolysis temperature of 550 degrees C with a sweeping gas flow rate of 100 cm(3) min(-1) in the presence of clinoptilolite (20% based on raw material) as catalyst. The pyrolytic and catalytic liquid products were analysed in detail to determine the predominant chemical classes and the identities of the major compounds present.     

Why Levallois? A Morphometric Comparison of Experimental ‘Preferential’ Levallois Flakes versus Debitage Flakes

Background

Middle Palaeolithic stone artefacts referred to as ‘Levallois’ have caused considerable debate regarding issues of technological predetermination, cognition and linguistic capacities in extinct hominins. Their association with both Neanderthals and early modern humans has, in particular, fuelled such debate. Yet, controversy exists regarding the extent of ‘predetermination’ and ‘standardization’ in so-called ‘preferential Levallois flakes’ (PLFs).

Methodology/Principal Findings

Using an experimental and morphometric approach, we assess the degree of standardization in PLFs compared to the flakes produced during their manufacture. PLFs possess specific properties that unite them robustly as a group or ‘category’ of flake. The properties that do so, relate most strongly to relative flake thicknesses across their surface area. PLFs also exhibit significantly less variability than the flakes generated during their production. Again, this is most evident in flake thickness variables. A further aim of our study was to assess whether the particular PLF attributes identified during our analyses can be related to current knowledge regarding flake functionality and utility.

Conclusions/Significance

PLFs are standardized in such a manner that they may be considered ‘predetermined’ with regard to a specific set of properties that distinguishes them statistically from a majority of other flakes. Moreover, their attributes can be linked to factors that, based on current knowledge, are desirable features in flake tools (e.g. durability, capacity for retouch, and reduction of torque). As such, our results support the hypothesis that the lengthy, multi-phase, and hierarchically organized process of Levallois reduction was a deliberate, engineered strategy orientated toward specific goals. In turn, our results support suggestions that Levallois knapping relied on a cognitive capacity for long-term working memory. This is consistent with recent evidence suggesting that cognitive distinctions between later Pleistocene hominins such as the Neanderthals and anatomically modern humans were not as sharp as some scholars have previously suggested.     

Structural basis for activation of an integral membrane protease by lipopolysaccharide

Omptins constitute a unique family of outer membrane proteases that are widespread in Enterobacteriaceae. The plasminogen activator (Pla) of Yersinia pestis is an omptin family member that is very important for development of both bubonic and pneumonic plague. The physiological function of Pla is to cleave (activate) human plasminogen to form the plasma protease plasmin. Uniquely, lipopolysaccharide (LPS) is essential for the catalytic activity of all omptins, including Pla. Why omptins require LPS for enzymatic activity is unknown. Here, we report the co-crystal structure of LPS-free Pla in complex with the activation loop peptide of human plasminogen, its natural substrate. The structure shows that in the absence of LPS, the peptide substrate binds deep within the active site groove and displaces the nucleophilic water molecule, providing an explanation for the dependence of omptins on LPS for enzymatic activity.     

Histological changes of liver glycogen storage in mice (Mus musculus) caused by high-protein diets

High Protein diets (HP) have been popular for people who want to lose weight since the 1960s. Even though these diets do not harm healthy people in the short term, there is insufficient data to support their safe use and efficiency over a long period. Because of the fact that the proteins in these diets are mainly from animal sources, it induces a higher intake of total fat, saturated fat and cholesterol. It is proven that high protein diets cause both physical and pathological abnormalities in the body. However, there exist very few studies about the effects of high protein nutrition on liver glycogen storage. For this study 40 Swiss albino mice consisting of two groups were used. The first group was fed with 25% High Protein; the other was fed with standard meal. The two groups were fed with respect to their diets for 30 days. At the end of 15th, 20th, 25th and 30th days 5 from each group were killed with cervical dislocation. The livers were removed after perfusion then fixated. The routine paraffin pursuit was applied before cutting into 5 micrometer sections and staining with H-E, PAS and silver. There were major differences in weight loss between the first and the fifteenth days. Glycogen storage was significantly reduced in HP (15) stained with PAS. Hydropic degeneration and regenerative activity was observed in H-E and silver stained HP group. As a result for the high protein diet group, weight loss at the 15th day and a significant decrement in glycogen storage at the 30th day was observed.     

Left ventricular giant pseudoaneurysm

Left ventricular (LV) pseudoaneurysm is a rare complication of myocardial infarction. It may also occur as a complication of mitral valve surgery, chest trauma, and bacterial endocarditis. It forms when a cardiac rupture contains adherent pericardium or scar tissue and is typically located on the posterior or inferior LV wall. Pseudoaneurysms have a propensity to spontaneous rupture; hence, immediate surgical intervention is the treatment of choice for LV pseudoaneurysms diagnosed in the first months after myocardial infarction. The management of chronic LV pseudoaneurysms is still a subject of debate.     

Influence of Dietary Copper Proteinate on Performance,Selected Biochemical Parameters,Lipid Peroxidation,Liver, and Egg Copper Content in Laying Hens

This study was performed to determine the effects of copper proteinate on performance, blood chemistry, lipid peroxidation status, and organs as well as copper deposition in the liver and eggs of laying hens. Seventy-two 30-week-old Bovans laying hens were distributed into four groups with three replicates. Animals were fed basal diet containing at least 17% crude protein and 2,800 kcal/kg metabolizable energy supplemented with either 0, 150, 300, or 450 mg/kg copper as copper proteinate. Supplementation of 150 and 300 mg/kg copper increased egg production, whereas 450 mg/kg copper decreased (p < 0.001). Liver copper levels were elevated in 300 and 450 mg/kg copper-supplemented groups (p < 0.001). Egg copper contents increased in all treatment groups (p < 0.01). An increase in glucose (p < 0.001) and decreases in albumin (p < 0.01) and total cholesterol (p < 0.05) levels were determined with 300 and 450 mg/kg copper. Supplementation of 450 mg/kg copper increased alkaline phosphatase and gamma glutamyl transpeptidase activities (p < 0.05), malondialdehyde, and high-density lipoprotein levels (p < 0.01) but decreased alanine aminotransferase and lactate dehydrogenase activities (p < 0.01). No gross and microscopic changes were observed in the liver and kidneys. These results indicated that 150 and 300 mg/kg copper increased egg production without having marked adverse effects, but 450 mg/kg copper altered some blood chemistry variables and reduced egg production in laying hens.     

Concentrated HCl Catalyzed 5-(Chloromethyl)furfural Production from Corn Stover of Varying Particle Sizes

5-(Chloromethyl) Furfural (CMF) is a potential chemical building block for replacing petroleum-derived chemicals derived from lignocellulosic feedstocks. In this study, hand harvested corn stover and mechanically forage chopped corn stover was processed in a 1 L hydrolysis reactor to produce CMF in a biphasic, two solvent system. Both 1,2 dichloroethane (DCE) and dichloromethane (DCM) were tested as organic solvents. The results showed that DCE performed better than DCM due to temperature and pressure limitations of the reactor system. Using DCE as the extracting solvent, the effects of solids loading, particle size, and moisture content of the corn stover on the hydrolysis efficiency were determined. One liter acid hydrolysis reactor provides consistent and reproducible yields of 63% CMF from hand harvested corn stover as feedstock at solid loading of 10% wt/v, 100C for 1 h. For the forage chopped corn stover, increasing particle size brings an increase in the feedstock sugar content. Foraged chopped corn stover (FCCS) particle sizes larger than 19 mm (0.75 in.) results in significant reduction in CMF yield from 43 to 35%.