Identification of double bond position in lipids : From GC to OzID

Recent developments in mass spectrometry and chromatography provide new possibilities for the identification and in some instances quantification of a wide range of lipids in complex matrices. These advances in analytical technologies have provided a tantalizing glimpse of the true structural diversity of lipids in nature and have reinvigorated interest in the role of lipids in biology. While technological advances have been impressive, difficulties in the ready identification of sites of unsaturation (i.e., double bond position) within these molecules presents a significant impediment to understanding lipid biochemistry. This is of particular importance given the growing body of literature suggesting that the presence of naturally occurring lipid double bond isomers can have a significant influence, both positive and negative, on the development of pathologies such as cancer, cardiovascular disease and type 2 diabetes. This article provides a critical review of the Current suite of analytical approaches to the challenge of identification of the position of carbon-carbon double bonds in intact lipids. Crown Copyright (C) 2009 Published by Elsevier B.V. All rights reserved.

Selective involvement of TIMP-2 in the second activational cleavage of pro-MMP-2 : refinement of the pro-MMP-2 activation mechanism

A tissue inhibitor of metalloproteinases-2 (TIMP-2)-independent mechanism for generating the first activational cleavage of pro-matrix metalloproteinase-2 (MMP-2) was identified in membrane type-1 MMP (MT1-MMP)-transfected MCF-7 cells and confirmed in TIMP-2-deficient fibroblasts. In contrast, the second MMP-2-activational step was found to be TIMP-2 dependent in both systems. MMP-2 hemopexin C-terminal domain was found to be critical for the first step processing, confirming a need for membrane tethering. We propose that the intermediate species of MMP-2 forms the well-established trimolecular complex (MT1-MMP/TIMP-2/MMP-2) for further TIMP-2-dependent autocatalytic cleavage to the fully active species. This alternate mechanism may supplement the traditional TIMP-2-mediated first step mechanism.

The role of the yeast cleavage and polyadenylation factor subunit Ydh1p/Cft2p in pre-mRNA 3'-end formation

Cleavage and polyadenylation factor (CPF) is a multi‐protein complex that functions in pre‐mRNA 3′‐end formation and in the RNA polymerase II (RNAP II) transcription cycle. Ydh1p/Cft2p is an essential component of CPF but its precise role in 3′‐end processing remained unclear. We found that mutations in YDH1 inhibited both the cleavage and the polyadenylation steps of the 3′‐end formation reaction in vitro. Recently, we demonstrated that an important function of CPF lies in the recognition of poly(A) site sequences and RNA binding analyses suggesting that Ydh1p/Cft2p interacts with the poly(A) site region. Here we show that mutant ydh1 strains are deficient in the recognition of the ACT1 cleavage site in vivo. The C‐terminal domain (CTD) of RNAP II plays a major role in coupling 3′‐end processing and transcription. We provide evidence that Ydh1p/Cft2p interacts with the CTD of RNAP II, several other subunits of CPF and with Pcf11p, a component of CF IA. We propose that Ydh1p/Cft2p contributes to the formation of important interaction surfaces that mediate the dynamic association of CPF with RNAP II, the recognition of poly(A) site sequences and the assembly of the polyadenylation machinery on the RNA substrate.

Characterization of two homologous disulfide bond systems involved in virulence factor Biogenesis in Uropathogenic Escherichia coli CFT073

Disulfide bond (DSB) formation is catalyzed by disulfide bond proteins and is critical for the proper folding and functioning of secreted and membrane-associated bacterial proteins. Uropathogenic Escherichia coli (UPEC) strains possess two paralogous disulfide bond systems: the well-characterized DsbAB system and the recently described DsbLI system. In the DsbAB system, the highly oxidizing DsbA protein introduces disulfide bonds into unfolded polypeptides by donating its redox-active disulfide and is in turn reoxidized by DsbB. DsbA has broad substrate specificity and reacts readily with reduced unfolded proteins entering the periplasm. The DsbLI system also comprises a functional redox pair; however, DsbL catalyzes the specific oxidative folding of the large periplasmic enzyme arylsulfate sulfotransferase (ASST). In this study, we characterized the DsbLI system of the prototypic UPEC strain CFT073 and examined the contributions of the DsbAB and DsbLI systems to the production of functional flagella as well as type 1 and P fimbriae. The DsbLI system was able to catalyze disulfide bond formation in several well-defined DsbA targets when provided in trans on a multicopy plasmid. In a mouse urinary tract infection model, the isogenic dsbAB deletion mutant of CFT073 was severely attenuated, while deletion of dsbLI or assT did not affect colonization.

Characterisation of thin layer polymer cement mortared concrete masonry bond

Masonry bond is affected by many parameters such as the type of mortar used, the techniques of dispersion of mortar and the surface texture of the concrete blocks. Additionally it is understood from the studies on conventional masonry that the bond characteristics are also influenced by the curing methods as well as the age of the bond at the time of testing. These effects on thin layer mortared masonry employing polymer cement mortars are not well understood. Therefore, the effect of curing methods and age to the bond strength and deformation of masonry containing thin layered polymer cement mortar was investigated as part of an ongoing research program at the Queensland University of Technology. This paper presents an experimental investigation of the flexural and shear bond characteristics of the thin layer mortared concrete masonry. The parameters examined include the effects curing and ageing to the bond development over a period from 14 days to 56 days after fabrication. The results exhibit that dry cured thin layer mortared masonry exhibits higher bond strength and Young’s and shear moduli compared to the wet cured specimens.

Effect of bond length on the behaviour of CFRP strengthened concrete-filled steel tubes under transverse impact

Concrete-filled steel tubular (CFST) columns have shown great potential as axial load carrying member and used widely in many mission critical infrastructures. However, attention is needed to strengthen these members where transverse impact force is expected to occur due to vehicle collisions. In this work, finite element (FE) model of carbon fibre reinforced polymer (CFRP) strengthened CFST columns are developed and the effect of CFRP bond length is investigated under transverse impact loading. Initially the numerical models have been validated by comparing impact test results from literature. The validated models are then used for detail parametric studies by varying the length of externally bonded CFRP composites. The parameters considered for this research are impact velocity, impact mass, CFRP modulus, adhesive type, and axial static loading. It has been observed that the effect of CFRP strengthening is consistent after an optimum effective bond length of CFRP wrapping. The effect of effective bond length has been studied for above parameters. The results show that, under combined axial static and transverse impact loads CFST columns can successfully prevent global buckling failure by strengthening only 34% of column length. Therefore, estimation of effective bond length is essential to utilise the CFRP composites cost effectively.

COL1A1 C-propeptide cleavage site mutation causes high bone mass, bone fragility and jaw lesions: A new cause of gnathodiaphyseal dysplasia?

Gnathodiaphyseal dysplasia (GDD) is a rare autosomal dominant condition characterized by bone fragility, irregular bone mineral density (BMD) and fibro-osseous lesions in the skull and jaw. Mutations in Anoctamin-5 (ANO5) have been identified in some cases. We aimed to identify the causative mutation in a family with features of GDD but no mutation in ANO5, using whole exome capture and massive parallel sequencing (WES). WES of two affected individuals (a mother and son) and the mother's unaffected parents identified a mutation in the C-propeptide cleavage site of COL1A1. Similar mutations have been reported in individuals with osteogenesis imperfecta (OI) and paradoxically increased BMD. C-propeptide cleavage site mutations in COL1A1 may not only cause 'high bone mass OI', but also the clinical features of GDD, specifically irregular sclerotic BMD and fibro-osseous lesions in the skull and jaw. GDD patients negative for ANO5 mutations should be assessed for mutations in type I collagen C-propeptide cleavage sites.

Bond Characteristics of Strengthened and Retrofitted Steel by Smart CFRP Technique

Usage of new smart materials in retrofitting of structures has become popular within last decade. Carbon fiber reinforced polymer (CFRP) has been widely used in retrofitting and strengthening of concrete structures and its usage in metallic structures is still in the developing stage. The variation of mechanical properties of CFRP and the consequent effects on strengthening and retrofitting CFRP systems are yet to be investigated under different loading and environmental conditions. This paper presents the results of CFRP strengthened and retrofitted corroded steel plate double strap joints under tension. An accelerated corrosion cell has been developed to accelerate the corrosion of the steel samples and CFRP strengthened samples. The results show a direct comparison of bond characteristics of CFRP strengthened and retrofitted steel double strap joints.