Albumin-mediated extracellular zinc speciation drives cellular zinc uptake

This work was financially supported by the Leverhulme Trust (RPG-2017-214) and BBSRC (BB/J006467/1 and BB/V014684/1). We thank Prof. Andrew Riches (University of St. Andrews) for provision of materials, and Dr. Elizabeth Bolitho (University of Warwick) for assistance with cell culture experiments.The role of the extracellular medium in influencing metal uptake into cells has not been described quantitatively. In a chemically defined model system containing albumin, zinc influx into endothelial cells correlates with the extracellular free zinc concentration. Allosteric inhibition of zinc-binding to albumin by free fatty acids increased zinc flux.Publisher PDFPeer reviewe

Ischemia-modified albumin : crosstalk between fatty acid and cobalt binding

This work was supported by the Leverhulme Trust (grant ref. RPG-2017-214), BBSRC (grant ref. BB/J006467/1) and the British Heart Foundation (grant refs. PG/15/9/31270 and FS/15/42/31556).Myocardial ischemia is difficult to diagnose effectively with still few well-defined biochemical markers for identification in advance, or in the absence of myocardial necrosis. “Ischemia-modified albumin” (IMA), a form of albumin displaying reduced cobalt-binding affinity, is significantly elevated in ischemic patients, and the albumin cobalt-binding (ACB) assay can measure its level indirectly. Elucidating the molecular mechanism underlying the identity of IMA and the ACB assay hinges on understanding metal-binding properties of albumin. Albumin binds most metal ions and harbours four primary metal binding sites: site A, site B, the N-terminal site (NTS), and the free thiol at Cys34. Previous efforts to clarify the identity of IMA and the causes for its reduced cobalt-binding capacity were focused on the NTS site, but the degree of N-terminal modification could not be correlated to the presence of ischemia. More recent work suggested that Co2+ ions as used in the ACB assay bind preferentially to site B, then to site A, and finally to the NTS. This insight paved the way for a new consistent molecular basis of the ACB assay: albumin is also the main plasma carrier for free fatty acids (FFAs), and binding of a fatty acid to the high-affinity site FA2 results in conformational changes in albumin which prevent metal binding at site A and partially at site B. Thus, this review advances the hypothesis that high IMA levels in myocardial ischemia and many other conditions originate from high plasma FFA levels hampering the binding of Co2+ to sites A and/or B. This is supported by biophysical studies and the co-association of a range of pathological conditions with positive ACB assays and high plasma FFA levels.Publisher PDFPeer reviewe

A single sensor controls large variations in zinc quotas in a marine cyanobacterium

Marine cyanobacteria are critical players in global nutrient cycles that crucially depend on trace metals in metalloenzymes, including zinc for CO2 fixation and phosphorus acquisition. How strains proliferating in the vast oligotrophic ocean gyres thrive at ultra-low zinc concentrations is currently unknown. Using Synechococcus sp. WH8102 as a model we show that its zinc-sensor protein Zur differs from all other known bacterial Zur proteins in overall structure and the location of its sensory zinc site. Uniquely, Synechococcus Zur activates metallothionein gene expression, which supports cellular zinc quotas spanning two orders of magnitude. Thus, a single zinc sensor facilitates growth across pico- to micromolar zinc concentrations with the bonus of banking this precious resource. The resultant ability to grow well at both ultra-low and excess zinc, together with overall lower zinc requirements, likely contribute to the broad ecological distribution of Synechococcus across the global oceans

Direct peptide bioconjugation/PEGylation at tyrosine with linear and branched polymeric diazonium salts

Direct polymer conjugation at peptide tyrosine residues is described. In this study Tyr residues of both leucine enkephalin and salmon calcitonin (sCT) were targeted using appropriate diazonium salt-terminated linear monomethoxy poly(ethylene glycol)s (mPEGs) and poly(mPEG) methacrylate prepared by atom transfer radical polymerization. Judicious choice of the reaction conditions-pH, stoichiometry, and chemical structure of diazonium salt-led to a high degree of site-specificity in the conjugation reaction, even in the presence of competitive peptide amino acid targets such as histidine, lysines, and N-terminal amine. In vitro studies showed that conjugation of mPEG 2000 to sCT did not affect the peptide's ability to increase intracellular cAMP induced in T47D human breast cancer cells bearing sCT receptors. Preliminary in vivo investigation showed preserved ability to reduce [Ca 2+] plasma levels by mPEG 2000-sCT conjugate in rat animal models. © 2012 American Chemical Society

Phosphorescent Sensor for Robust Quantification of Copper(II) Ion

A phosphorescent sensor based on a multichromophoric iridium(III) complex was synthesized and characterized. The construct exhibits concomitant changes in its phosphorescence intensity ratio and phosphorescence lifetime in response to copper(II) ion. The sensor, which is reversible and selective, is able to quantify copper(II) ions in aqueous media, and it detects intracellular copper ratiometrically.National Institute of General Medical Sciences (U.S.) ((Grant GM065519)Ewha Woman's University (Korea) (RP-Grant 2009

Comparative genomics of Pseudomonas fluorescens subclade III strains from human lungs

Abstract Background While the taxonomy and genomics of environmental strains from the P. fluorescens species-complex has been reported, little is known about P. fluorescens strains from clinical samples. In this report, we provide the first genomic analysis of P. fluorescens strains in which human vs. environmental isolates are compared. Results Seven P. fluorescens strains were isolated from respiratory samples from cystic fibrosis (CF) patients. The clinical strains could grow at a higher temperature (>34 °C) than has been reported for environmental strains. Draft genomes were generated for all of the clinical strains, and multi-locus sequence analysis placed them within subclade III of the P. fluorescens species-complex. All strains encoded type- II, −III, −IV, and -VI secretion systems, as well as the widespread colonization island (WCI). This is the first description of a WCI in P. fluorescens strains. All strains also encoded a complete I2/PfiT locus and showed evidence of horizontal gene transfer. The clinical strains were found to differ from the environmental strains in the number of genes involved in metal resistance, which may be a possible adaptation to chronic antibiotic exposure in the CF lung. Conclusions This is the largest comparative genomics analysis of P. fluorescens subclade III strains to date and includes the first clinical isolates. At a global level, the clinical P. fluorescens subclade III strains were largely indistinguishable from environmental P. fluorescens subclade III strains, supporting the idea that identifying strains as ‘environmental’ vs ‘clinical’ is not a phenotypic trait. Rather, strains within P. fluorescens subclade III will colonize and persist in any niche that provides the requirements necessary for growth.http://deepblue.lib.umich.edu/bitstream/2027.42/116129/1/12864_2015_Article_2261.pd