A molecular dynamics study based post facto free energy analysis of the binding of bovine angiogenin with UMP and CMP ligands

Angiogenin is a protein belonging to the superfamily of RNase A. The RNase activity of this protein is essential for its angiogenic activity. Although members of the RNase A family carry out RNase activity, they differ markedly in their strength and specificity. In this paper, we address the problem of higher specificity of angiogenin towards cytosine against uracil in the first base binding position. We have carried out extensive nano-second level molecular dynamics(MD) computer simulations on the native bovine angiogenin and on the CMP and UMP complexes of this protein in aqueous medium with explicit molecular solvent. The structures thus generated were subjected to a rigorous free energy component analysis to arrive at a plausible molecular thermodynamic explanation for the substrate specificity of angiogenin.

Efecto de diferentes dosis de nitrógeno y del inhibidor CMP sobre el comportamiento del arroz (Oryza sativa L.) var. Altamira-7 y la dinámica de las malezas

El presente trabajo se realizó en el Centro Experimental El Zonajel en el municipio de San Lorenzo Boaco, V Región; con la finalidad de determinar el efecto de diferentes dosis de nitrógeno y del inhibidor CMP ( 1 Carbamyl, 3-5 methyl pirazole) sobre el comportamiento del Arroz (Oryza sativa , L) variedad “Altamira 7” y la dinámica de las malezas; el periodo de evaluación se llevó a cabo de Septiembre de 1989 a Enero de 1990 en un diseño de bloques completos al azar con 14 tratamientos (15N +30N, 30N+60Nm 45N 90N, 22.5N+22.5, 45N+45N, 45N, 90N, 60N+6CMP, 80N+6CMO. 60N+8CMP, 80N +8CMP. 0N y 8CMP kg./ha.) bajo aniego. Los resultados indican que los tratamientos con el inhibidor CMP reducen la abundancia de malezas. Aunque estos a la vez aumentan la dominancia de las mismas. Altamira 7 respondió significativamente a la aplicación de nitrógeno dado que se notaron incrementos en la altura, ahijamiento y biomasa del cultivo, no encontrándose diferencias significativas entre los tratamientos con fertilizantes nitrogenados. Los resultados similares se obtuvieron en población de panícula y longitud de panícula: En cuanto al peso de mil semillas no hubo diferencias marcadas entre los tratamientos. Respecto al rendimiento quedo demostrado que las dosis de fertilizantes nitrogenados entre 60y 80 kg/ha responden a las necesidades de este nutriente para Altamira 7.

CMP流场的数值模拟及离心力影响分析

化学机械抛光(chemical mechanical polishing,CMP)是一项融合化学分解和机械力学的工艺,其中包含了流体动力润滑的作用.在已有润滑方程的基础上,提出并分析了带有离心力项的润滑方程.利用Chebyshev加速超松弛技术对有离心力项的润滑方程进行求解,得到离心力对抛光液压力分布的影响.数值模拟结果表明,压力分布与不带离心力项的润滑方程得出的明显不同;无量纲载荷和转矩随中心膜厚、转角、倾角、抛光垫旋转角速度等参数的变化趋势相同,但数值相差较大,抛光垫旋转角速度越大差别越大.

CMP润滑方程的Chebyshev加速超松弛法求解

化学机械抛光（chemicalmechanicalpolishing，CMP）是一项融合了化学分解和机械力学的工艺技术，其中包含了流体动力润滑的作用．文中通过分析流场效应，建立了Sum在化学机械抛光实验基础上获得的润滑（Reynolds）方程，利用Chebyshev加速超松弛技术对润滑方程进行求解，得到了新的抛光液压力分布模型，给出了一些新的载荷及转矩在不同参数下的变化情况．

Over Expression of the CMP-sialic Acid Transporter in Chinese Hamster Ovary Cells Leads to Increased Sialylation

Most glyco-engineering approaches used to improve quality of recombinant glycoproteins involve the manipulation of glycosyltransferase and/or glycosidase expression. We investigated whether the over expression of nucleotide sugar transporters, particularly the CMP-sialic acid transporter (CMP-SAT), would be a means to improve the sialylation process in CHO cells. We hypothesized that increasing the expression of the CMP-SAT in the cells would increase the transport of the CMP-sialic acid in the Golgi lumen, hence increasing the intra-lumenal CMP-sialic acid pool, and resulting in a possible increase in sialylation extent of proteins being produced. We report the construction of a CMP-SAT expression vector which was used for transfection into CHO-IFNγ, a CHO cell line producing human IFNγ. This resulted in approximately 2 to 5 times increase in total CMP-SAT expression in some of the positive clones as compared to untransfected CHO-IFNγ, as determined using real-time PCR analysis. This in turn concurred with a 9.6% to 16.3% percent increase in site sialylation. This engineering approach has thus been identified as a novel means of improving sialylation in recombinant glycoprotein therapeutics. This strategy can be utilized feasibly on its own, or in combination with existing sialylation improvement strategies. It is believed that such multi-prong approaches are required to effectively manipulate the complex sialylation process, so as to bring us closer to the goal of producing recombinant glycoproteins of high and consistent sialylation from mammalian cells.

The Rv1712 locus from Mycobacterium tuberculosis H37Rv codes for a functional CMP kinase that preferentially Phosphorylates dCMP

The Mycobacterium tuberculosis cmk gene, predicted to encode a CMP kinase (CMK), was cloned and expressed, and its product was purified to homogeneity. Steady-state kinetics confirmed that M. tuberculosis CMK is a monomer that preferentially phosphorylates CMP and dCMP by a sequential mechanism. A plausible role for CMK is discussed. Copyright © 2009, American Society for Microbiology. All Rights Reserved.

Determinação de GMP e CMP* no leite por métodos espectrofotométrico (ANSM) e cromatográfico (HPLC) - parâmetros metodológicos (*glicomacropeptídeo e caseinomacropeptídeo)

Glycomacropeptide is a glycosilated fraction of bovine kappa-casein that remains soluble when milk is clotted by rennin. Determinations of milk sialic acid content are useful because its concentration reflects the amount of free GMP of milk. In normal milk these amounts are very low, 12 to 16 times lower than in sweet whey. Therefore, its determination may be applied to verify possible frauds with whey addictions, since it works as a fingerprint. With the description of a new spectrophotometric method for determination of free GMP (ANSM) occurred a simplification of procedures, being faster than others (HPLC method), without loss of accuracy. However, due to variations of glycosilation in kappa-casein between animals, during the lactation period, due to mastitis and yet due to proteolysis on milk, it was necessary to know these variations to interpret correctly the analytical results. It was analyzed 1,703 samples of producer's raw milk and 1,189 samples of processed milk (HTST and UHT). The results showed that normal milk from herd (producer's milk) have only small amounts of free GMP, with A470nm = 0.232±0.088 or 3.89±1.25 mg of sialic acid/L. The upper limit of this distribution was A = 0.496; thus every bigger value may represent a problem, being outside of normal distribution.

Cloning, Expression in Escherichia coli, and Characterization of Arabidopsis thaliana UMP/CMP Kinase1

A cDNA encoding the Arabidopsis thaliana uridine 5′-monophosphate (UMP)/cytidine 5′-monophosphate (CMP) kinase was isolated by complementation of a Saccharomyces cerevisiae ura6 mutant. The deduced amino acid sequence of the plant UMP/CMP kinase has 50% identity with other eukaryotic UMP/CMP kinase proteins. The cDNA was subcloned into pGEX-4T-3 and expressed as a glutathione S-transferase fusion protein in Escherichia coli. Following proteolytic digestion, the plant UMP/CMP kinase was purified and analyzed for its structural and kinetic properties. The mass, N-terminal sequence, and total amino acid composition agreed with the sequence and composition predicted from the cDNA sequence. Kinetic analysis revealed that the UMP/CMP kinase preferentially uses ATP (Michaelis constant [Km] = 29 μm when UMP is the other substrate and Km = 292 μm when CMP is the other substrate) as a phosphate donor. However, both UMP (Km = 153 μm) and CMP (Km = 266 μm) were equally acceptable as the phosphate acceptor. The optimal pH for the enzyme is 6.5. P1, P5-di(adenosine-5′) pentaphosphate was found to be a competitive inhibitor of both ATP and UMP.

Expression cloning of the Golgi CMP-sialic acid transporter.

Translocation of nucleotide sugars across the membrane of the Golgi apparatus is a prerequisite for the synthesis of complex carbohydrate structures. While specific transport systems for different nucleotide sugars have been identified biochemically in isolated microsomes and Golgi vesicles, none of these transport proteins has been characterized at the molecular level. Chinese hamster ovary (CHO) mutants of the complementation group Lec2 exhibit a strong reduction in sialylation of glycoproteins and glycolipids due to a defect in the CMP-sialic acid transport system. By complementation cloning in the mutant 6B2, belonging to the Lec2 complementation group, we were able to isolate a cDNA encoding the putative murine Golgi CMP-sialic acid transporter. The cloned cDNA encodes a highly hydrophobic, multiple membrane spanning protein of 36.4 kDa, with structural similarity to the recently cloned ammonium transporters. Transfection of a hemagglutinin-tagged fusion protein into the mutant 6B2 led to Golgi localization of the hemagglutinin epitope. Our results, together with the observation that the cloned gene shares structural similarities to other recently cloned transporter proteins, strongly suggest that the isolated cDNA encodes the CMP-sialic acid transporter.

Saving schools from abomination and abnormal sex : a discourse analysis of online public commentary about 'queering' school spaces

Homophobic hatred: these words summarise online commentary made by people in support of a school that banned gay students from taking their same sex partners to a school formal. With the growing popularity of online news sites, it seems appropriate to critically examine how these sites are becoming a new arena in which people can express personal opinions about controversial topics. While commentators equally expressed two dominant viewpoints about the school ban (homophobic hatred and human rights), this paper focuses on homophobic hatred as a discursive position and how the comments work to confirm the legitimacy of the schools’ decision. Drawing on the work of Foucault and others, the paper examines how the comments constitute certain types of subjectivity drawing on dominant ideas about what it means to be homophobic. The analysis demonstrates the complex and competing skein of strategies that constitute queering school social spaces as a social problem.