Carboxysomal Carbonic Anhydrases: Structure and Role in Microbial CO\u3csub\u3e2\u3c/sub\u3e Fixation

Cyanobacteria and some chemoautotrophic bacteria are able to grow in environments with limiting CO2 concentrations by employing a CO2-concentrating mechanism (CCM) that allows them to accumulate inorganic carbon in their cytoplasm to concentrations several orders of magnitude higher than that on the outside. The final step of this process takes place in polyhedral protein microcompartments known as carboxysomes, which contain the majority of the CO2-fixing enzyme, RubisCO. The efficiency of CO2 fixation by the sequestered RubisCO is enhanced by co-localization with a specialized carbonic anhydrase that catalyzes dehydration of the cytoplasmic bicarbonate and ensures saturation of RubisCO with its substrate, CO2. There are two genetically distinct carboxysome types that differ in their protein composition and in the carbonic anhydrase(s) they employ. Here we review the existing information concerning the genomics, structure and enzymology of these uniquely adapted carbonic anhydrases, which are of fundamental importance in the global carbon cycle

Partial purification and characterization of the DNA polymerase from the cyanelles of Cyanophora paradoxa

AbstractA DNA polymerase was partially purified and characterized from the photosynthetic organelles (cyanelles) of the protist, Cyanophora paradoxa. While cyanelles have several cyanobacterial features, such as a lysozyme-sensitive cell wall, unstacked thylakoids and light harvesting phycobilisomes, their genome size and structure resemble those of chloroplasts, suggesting that cyanelles occupy a unique intermediate position between chloroplasts and their phylogenetic ancestors, the cyanobacteria. When comparing the biochemical characteristics of the cyanelle DNA polymerase to those of its counterparts from higher plant chloroplasts and from a cyanobacterium, it is clear that the cyanelle enzyme resembles chloroplast DNA polymerases which are eukaryotic γ-type enzymes

The Structure of β-Carbonic Anhydrase from the Carboxysomal Shell Reveals a Distinct Subclass with One Active Site for the Price of Two

CsoSCA (formerly CsoS3) is a bacterial carbonic anhydrase localized in the shell of a cellular microcompartment called the carboxysome, where it converts HCO-3 to CO2 for use in carbon fixation by ribulose-bisphosphate carboxylase/oxygenase (RuBisCO). CsoSCA lacks significant sequence similarity to any of the four known classes of carbonic anhydrase (α, β, γ, or δ), and so it was initially classified as belonging to a new class, ϵ. The crystal structure of CsoSCA from Halothiobacillus neapolitanus reveals that it is actually a representative member of a new subclass of β-carbonic anhydrases, distinguished by a lack of active site pairing. Whereas a typical β-carbonic anhydrase maintains a pair of active sites organized within a two-fold symmetric homodimer or pair of fused, homologous domains, the two domains in CsoSCA have diverged to the point that only one domain in the pair retains a viable active site. We suggest that this defunct and somewhat diminished domain has evolved a new function, specific to its carboxysomal environment. Despite the level of sequence divergence that separates CsoSCA from the other two subclasses of β-carbonic anhydrases, there is a remarkable level of structural similarity among active site regions, which suggests a common catalytic mechanism for the interconversion of HCO-3 and CO2. Crystal packing analysis suggests that CsoSCA exists within the carboxysome shell either as a homodimer or as extended filaments

Efficacy and safety of epratuzumab in patients with moderate/severe active systemic lupus erythematosus: results from EMBLEM, a phase IIb, randomised, double-blind, placebo-controlled, multicentre study

Objective: To identify a suitable dosing regimen of the CD22-targeted monoclonal antibody epratuzumab in adults with moderately to severely active systemic lupus erythematosus (SLE). Methods: A phase IIb, multicentre, randomised controlled study (NCT00624351) was conducted with 227 patients (37–39 per arm) receiving either: placebo, epratuzumab 200 mg cumulative dose (cd) (100 mg every other week (EOW)), 800 mg cd (400 mg EOW), 2400 mg cd (600 mg weekly), 2400 mg cd (1200 mg EOW), or 3600 mg cd (1800 mg EOW). The primary endpoint (not powered for significance) was the week 12 responder rate measured using a novel composite endpoint, the British Isles Lupus Assessment Group (BILAG)-based Combined Lupus Assessment (BICLA). Results: Proportion of responders was higher in all epratuzumab groups than with placebo (overall treatment effect test p=0.148). Exploratory pairwise analysis demonstrated clinical improvement in patients receiving a cd of 2400 mg epratuzumab (OR for 600 mg weekly vs placebo: 3.2 (95% CI 1.1 to 8.8), nominal p=0.03; OR for 1200 mg EOW vs placebo: 2.6 (0.9 to 7.1), nominal p=0.07). Post-hoc comparison of all 2400 mg cd patients versus placebo found an overall treatment effect (OR=2.9 (1.2 to 7.1), nominal p=0.02). Incidence of adverse events (AEs), serious AEs and infusion reactions was similar between epratuzumab and placebo groups, without decreases in immunoglobulin levels and only partial reduction in B-cell levels. Conclusions: Treatment with epratuzumab 2400 mg cd was well tolerated in patients with moderately to severely active SLE, and associated with improvements in disease activity. Phase III studies are ongoing

Structural Analysis of CsoS1A and the Protein Shell of the \u3ci\u3eHalothiobacillus neapolitanus\u3c/i\u3e Carboxysome

The carboxysome is a bacterial organelle that functions to enhance the efficiency of CO2 fixation by encapsulating the enzymes ribulose bisphosphate carboxylase/ oxygenase (RuBisCO) and carbonic anhydrase. The outer shell of the carboxysome is reminiscent of a viral capsid, being constructed from many copies of a few small proteins. Here we describe the structure of the shell protein CsoS1A from the chemoautotrophic bacterium Halothiobacillus neapolitanus. The CsoS1A protein forms hexameric units that pack tightly together to form a molecular layer, which is perforated by narrow pores. Sulfate ions, soaked into crystals of CsoS1A, are observed in the pores of the molecular layer, supporting the idea that the pores could be the conduit for negatively charged metabolites such as bicarbonate, which must cross the shell. The problem of diffusion across a semiporous protein shell is discussed, with the conclusion that the shell is sufficiently porous to allow adequate transport of small molecules. The molecular layer formed by CsoS1A is similar to the recently observed layers formed by cyanobacterial carboxysome shell proteins. This similarity supports the argument that the layers observed represent the natural structure of the facets of the carboxysome shell. Insights into carboxysome function are provided by comparisons of the carboxysome shell to viral capsids, and a comparison of its pores to the pores of transmembrane protein channels

Advances in Understanding Carboxysome Assembly in \u3ci\u3eProchlorococcus\u3c/i\u3e and \u3ci\u3eSynechococcus\u3c/i\u3e Implicate CsoS2 as a Critical Component

The marine Synechococcus and Prochlorococcus are the numerically dominant cyanobacteria in the ocean and important in global carbon fixation. They have evolved a CO2-concentrating-mechanism, of which the central component is the carboxysome, a self-assembling proteinaceous organelle. Two types of carboxysome, α and β, encapsulating form IA and form IB d-ribulose-1,5-bisphosphate carboxylase/oxygenase, respectively, differ in gene organization and associated proteins. In contrast to the β-carboxysome, the assembly process of the α-carboxysome is enigmatic. Moreover, an absolutely conserved α-carboxysome protein, CsoS2, is of unknown function and has proven recalcitrant to crystallization. Here, we present studies on the CsoS2 protein in three model organisms and show that CsoS2 is vital for α-carboxysome biogenesis. The primary structure of CsoS2 appears tripartite, composed of an N-terminal, middle (M)-, and C-terminal region. Repetitive motifs can be identified in the N- and M-regions. Multiple lines of evidence suggest CsoS2 is highly flexible, possibly an intrinsically disordered protein. Based on our results from bioinformatic, biophysical, genetic and biochemical approaches, including peptide array scanning for protein-protein interactions, we propose a model for CsoS2 function and its spatial location in the α-carboxysome. Analogies between the pathway for β-carboxysome biogenesis and our model for α-carboxysome assembly are discussed

Impacts of Climate Change on indirect human exposure to pathogens and chemicals from agriculture

Objective: Climate change is likely to affect the nature of pathogens and chemicals in the environment and their fate and transport. Future risks of pathogens and chemicals could therefore be very different from those of today. In this review, we assess the implications of climate change for changes in human exposures to pathogens and chemicals in agricultural systems in the United Kingdom and discuss the subsequent effects on health impacts. Data sources: In this review, we used expert input and considered literature on climate change ; health effects resulting from exposure to pathogens and chemicals arising from agriculture ; inputs of chemicals and pathogens to agricultural systems ; and human exposure pathways for pathogens and chemicals in agricultural systems. Data synthesis: We established the current evidence base for health effects of chemicals and pathogens in the agricultural environment ; determined the potential implications of climate change on chemical and pathogen inputs in agricultural systems ; and explored the effects of climate change on environmental transport and fate of different contaminant types. We combined these data to assess the implications of climate change in terms of indirect human exposure to pathogens and chemicals in agricultural systems. We then developed recommendations on future research and policy changes to manage any adverse increases in risks. Conclusions: Overall, climate change is likely to increase human exposures to agricultural contaminants. The magnitude of the increases will be highly dependent on the contaminant type. Risks from many pathogens and particulate and particle-associated contaminants could increase significantly. These increases in exposure can, however, be managed for the most part through targeted research and policy changes

Flow of bottom water in the northwestern Weddell Sea

The Weddell Sea is known to feed recently formed deep and bottom water into the Antarctic circumpolar water belt, from whence it spreads into the basins of the world ocean. The rates are still a matter of debate. To quantify the flow of bottom water in the northwestern Weddell Sea data obtained during five cruises with R/V Polarstern between October 1989 and May 1998 were used. During the cruises in the Weddell Sea, five hydrographic surveys were carried out to measure water mass properties, and moored instruments were deployed over a time period of 8.5 years to obtain quasi-continuous time series. The average flow in the bottom water plume in the northwestern Weddell Sea deduced from the combined conductivity-temperature-depth and moored observations is 1.3±0.4 Sv. Intensive fluctuations of a wide range of timescales including annual and interannual variations are superimposed. The variations are partly induced by fluctuations in the formation rates and partly by current velocity fluctuations related to the large-scale circulation. Taking into account entrainment of modified Warm Deep Water and Weddell Sea Deep Water during the descent of the plume along the slope, between 0.5 Sv and 1.3 Sv of surface-ventilated water is supplied to the deep sea. This is significantly less than the widely accepted ventilation rates of the deep sea. If there are no other significant sources of newly ventilated water in the Weddell Sea, either the dominant role of Weddell Sea Bottom Water in the Southern Ocean or the global ventilation rates have to be reconsidered