Separation Options for Phosphorylated Osteopontin from Transgenic Microalgae Chlamydomonas reinhardtii.

Correct folding and post-translational modifications are vital for therapeutic proteins to elicit their biological functions. Osteopontin (OPN), a bone regenerative protein present in a range of mammalian cells, is an acidic phosphoprotein with multiple potential phosphorylation sites. In this study, the ability of unicellular microalgae, Chlamydomonas reinhardtii, to produce phosphorylated recombinant OPN in its chloroplast is investigated. This study further explores the impact of phosphorylation and expression from a "plant-like" algae on separation of OPN. Chromatography resins ceramic hydroxyapatite (CHT) and Gallium-immobilized metal affinity chromatography (Ga-IMAC) were assessed for their binding specificity to phosphoproteins. Non-phosphorylated recombinant OPN expressed in E. coli was used to compare the specificity of interaction of the resins to phosphorylated OPN. We observed that CHT binds OPN by multimodal interactions and was better able to distinguish phosphorylated proteins in the presence of 250 mM NaCl. Ga-IMAC interaction with OPN was not selective to phosphorylation, irrespective of salt, as the resin bound OPN from both algal and bacterial sources. Anion exchange chromatography proved an efficient capture method to partially separate major phosphorylated host cell protein impurities such as Rubisco from OPN

Symmetries, Currents and Conservation Laws of Self-Dual Gravity

We describe an infinite-dimensional algebra of hidden symmetries for the self-dual gravity equations. Besides the known diffeomorphism-type symmetries (affine extension of w(infinity) algebra), this algebra contains new hidden symmetries, which are an affine extension of the Lorentz rotations. The full symmetry algebra has both Kac-Moody and Virasoro-like generators, whose exponentiation maps solutions of the field equations to other solutions. Relations to problems of string theories are briefly discussed.Comment: 14 pages, LaTeX, the paper was reformatte

Assessment of predation by the West Coast rock lobster (Jasus lalandii) : relationships among growth rate, diet and benthic community composition, with implications for the survival of juvenile abalone (Haliotis midae)

Bibliography: leaves 195-213.This thesis focused on two aspects of predation by the West Coast rock lobster (Jasus lalandii). These were (1) the problem of slow growth currently observed in adult male rock lobsters and the putative link between growth rate and food availability, and (2) the influence of rock-lobster predation on the benthos, but more specifically on sea urchins and juvenile abalone

The \u3cem\u3eChlamydomonas\u3c/em\u3e Genome Reveals the Evolution of Key Animal and Plant Functions

Chlamydomonas reinhardtii is a unicellular green alga whose lineage diverged from land plants over 1 billion years ago. It is a model system for studying chloroplast-based photosynthesis, as well as the structure, assembly, and function of eukaryotic flagella (cilia), which were inherited from the common ancestor of plants and animals, but lost in land plants. We sequenced the ∼120-megabase nuclear genome of Chlamydomonas and performed comparative phylogenomic analyses, identifying genes encoding uncharacterized proteins that are likely associated with the function and biogenesis of chloroplasts or eukaryotic flagella. Analyses of the Chlamydomonas genome advance our understanding of the ancestral eukaryotic cell, reveal previously unknown genes associated with photosynthetic and flagellar functions, and establish links between ciliopathy and the composition and function of flagella

Predicting biological variation using a simple morphometric marker in the sedentary marine invertebrate Haliotis rubra

Copyright © Inter-Research 2008Many sedentary marine invertebrates have a fine-scale (100s m) population structure that complicates their conservation and management. This is a consequence of the limited information on the boundaries between component populations and the biological variability among them. Blacklip abalone Haliotis rubra form discrete populations, many of which are ‘stunted’ with individuals reaching a maximum length less than those in adjacent areas. In the present study, we obtained information on the growth, size at maturity and fecundity of H. rubra from stunted and ‘non-stunted’ populations spread across broad (10s km) and fine (100s m) spatial scales. Relationships between each of these key population parameters and a simple ‘morphometric marker’ based on the relationship between shell length and shell height were also examined. Variation in broad-scale growth and size at maturity could primarily be attributed to differences between stunted and non-stunted sites. Within the stunted site, growth and size at maturity were substantially different over distances >150 m. However, within the non-stunted site these parameters tended to be similar across 1000 m. While the lowest fecundities tended to be in the stunted sites, there was significant overlap among all sites. These spatial patterns in biology were highly correlated with the spatial variability observed in a simple morphometric marker. These results suggest that this morphometric marker can be used as a tool for the spatial management of abalone fisheries by cheaply inferring key biological parameters for individual populations and identifing the boundaries among these based on these differences.T. Saunders and S. Mayfiel

Development of a cell surface display system in Chlamydomonas reinhardtii

Cell-surface display systems are biotechnological techniques used to express heterologous proteins on the cell surface. Their application depends directly on the cell system used, as well as on the anchoring point for the surface displayed protein. To meet most application demands an inexpensive, safe, and scalable production platform, that reduces the economic barriers for large scale use is needed. Toward this goal, we screened three possible cell surface anchoring points in the green algae Chlamydomonas by fusing mVenus to prospective anchors moieties. The vectors harboring mVenus:anchor were screened for mVenus fluorescence and tested for cellular localization by confocal laser scanning microscopy. This strategy allowed the identification of two functional anchors, one for the cytoplasmic membrane using the MAW8 GPI-anchor signal, and one for the cell wall using the GP1 protein. We also exploited GP1 chemical and biological traits to release the fused proteins efficiently during cell wall shedding. Our work provides a foundation for surface engineering of C reinhardtii supporting both cell biology studies and biotechnology applications