Domain Swapping within PDZ2 Is Responsible for Dimerization of ZO Proteins

ZO-1 is a multidomain protein involved in cell-cell junctions and contains three PDZ domains, which are necessary for its function in vivo. PDZ domains play a central role in assembling diverse protein complexes through their ability to recognize short peptide motifs on other proteins. We determined the structure of the second of the three PDZ domains of ZO-1, which is known to promote dimerization as well as bind to C-terminal sequences on connexins. The dimer is stabilized by extensive symmetrical domain swapping of β-strands, which is unlike any other known mechanism of PDZ dimerization. The canonical peptide-binding groove remains intact in both subunits of the PDZ2 dimer and is created by elements contributed from both monomers. This unique structure reveals an additional example of how PDZ domains dimerize and has multiple implications for both peptide binding and oligomerization in vivo

The Src Homology 3 Domain Is Required for Junctional Adhesion Molecule Binding to the Third PDZ Domain of the Scaffolding Protein ZO-1

Tight junctions are cell-cell contacts that regulate the paracellular flux of solutes and prevent pathogen entry across cell layers. The assembly and permeability of this barrier are dependent on the zonula occludens (ZO) membrane-associated guanylate kinase (MAGUK) proteins ZO-1, -2, and -3. MAGUK proteins are characterized by a core motif of protein-binding domains that include a PDZ domain, a Src homology 3 (SH3) domain, and a region of homology to guanylate kinase (GUK); the structure of this core motif has never been determined for any MAGUK. To better understand how ZO proteins organize the assembly of protein complexes we have crystallized the entire PDZ3-SH3-GUK core motif of ZO-1. We have also crystallized this core motif in complex with the cytoplasmic tail of the ZO-1 PDZ3 ligand, junctional adhesion molecule A (JAM-A) to determine how the activity of different domains is coordinated. Our study shows a new feature for PDZ class II ligand binding that implicates the two highly conserved Phe−2 and Ser−3 residues of JAM. Our x-ray structures and NMR experiments also show for the first time a role for adjacent domains in the binding of ligands to PDZ domains in the MAGUK proteins family

Finishing the euchromatic sequence of the human genome

The sequence of the human genome encodes the genetic instructions for human physiology, as well as rich information about human evolution. In 2001, the International Human Genome Sequencing Consortium reported a draft sequence of the euchromatic portion of the human genome. Since then, the international collaboration has worked to convert this draft into a genome sequence with high accuracy and nearly complete coverage. Here, we report the result of this finishing process. The current genome sequence (Build 35) contains 2.85 billion nucleotides interrupted by only 341 gaps. It covers ∼99% of the euchromatic genome and is accurate to an error rate of ∼1 event per 100,000 bases. Many of the remaining euchromatic gaps are associated with segmental duplications and will require focused work with new methods. The near-complete sequence, the first for a vertebrate, greatly improves the precision of biological analyses of the human genome including studies of gene number, birth and death. Notably, the human enome seems to encode only 20,000-25,000 protein-coding genes. The genome sequence reported here should serve as a firm foundation for biomedical research in the decades ahead

Process framework for identifying and prioritizing water quality improvement for meetings TMDLs in Texas

Center for Research in Water Resource

Insights into Regulated Ligand Binding Sites from the Structure of ZO-1 Src Homology 3-Guanylate Kinase Module*

Tight junctions are dynamic components of epithelial and endothelial cells that regulate the paracellular transport of ions, solutes, and immune cells. The assembly and permeability of these junctions is dependent on the zonula occludens (ZO) proteins, members of the membrane-associated guanylate kinase homolog (MAGUK) protein family, which are characterized by a core Src homology 3 (SH3)-GUK module that coordinates multiple protein-protein interactions. The structure of the ZO-1 SH3-GUK domain confirms that the interdependent folding of the SH3 and GUK domains is a conserved feature of MAGUKs, but differences in the orientation of the GUK domains in three different MAGUKs reveal interdomain flexibility of the core unit. Using pull-down assays, we show that an effector loop, the U6 region in ZO-1, forms a novel intramolecular interaction with the core module. This interaction is divalent cation-dependent and overlaps with the binding site for the regulatory molecule calmodulin on the GUK domain. These findings provide insight into the previously observed ability of the U6 region to regulate TJ assembly in vivo and the structural basis for the complex protein interactions of the MAGUK family

Fourier Analysis for Demand Forecasting in a Fashion Company

In the fashion industry, demand forecasting is particularly complex: companies operate with a large variety of short lifecycle products, deeply influenced by seasonal sales, promotional events, weather conditions, advertising and marketing campaigns, on top of festivities and socio-economic factors. At the same time, shelf-out-of-stock phenomena must be avoided at all costs. Given the strong seasonal nature of the products that characterize the fashion sector, this paper aims to highlight how the Fourier method can represent an easy and more effective forecasting method compared to other widespread heuristics normally used. For this purpose, a comparison between the fast Fourier transform algorithm and another two techniques based on moving average and exponential smoothing was carried out on a set of 4-year historical sales data of a €60+ million turnover medium- to large-sized Italian fashion company, which operates in the women's textiles apparel and clothing sectors. The entire analysis was performed on a common spreadsheet, in order to demonstrate that accurate results exploiting advanced numerical computation techniques can be carried out without necessarily using expensive software

Structural requirements for calmodulin binding to membrane-associated guanylate kinase homologs

Effector molecules such as calmodulin modulate the interactions of membrane-associated guanylate kinase homologs (MAGUKs) and other scaffolding proteins of the membrane cytoskeleton by binding to the Src homology 3 (SH3) domain, the guanylate kinase (GK) domain, or the connecting HOOK region of MAGUKs. Using surface plasmon resonance, we studied the interaction of members of all four MAGUK subfamilies—synapse-associated protein 97 (SAP97), calcium/calmodulin-dependent serine protein kinase (CASK), membrane palmitoylated protein 2 (MPP2), and zona occludens (ZO) 1—and calmodulin to determine interaction affinities and localize the binding site. The SH3-GK domains of the proteins and derivatives thereof were expressed in E. coli and purified. In all four proteins, high-affinity calmodulin binding was identified. CASK was shown to contain a Ca2+-dependent calmodulin binding site within the HOOK region, overlapping with a protein 4.1 binding site. In ZO1, a Ca2+-dependent calmodulin binding site was detected within the GK domain. The equilibrium dissociation constants for MAGUK–calmodulin interaction were found to range from 50 nM to 180 nM. Sequence analyses suggest that binding sites for calmodulin have evolved independently in at least three subfamilies. For ZO1, pulldown of GST-calmodulin was shown to occur in a calcium-dependent manner; moreover, molecular modeling and sequence analyses predict conserved basic residues to be exposed on one side of a helix. Thus, calmodulin binding appears to be a common feature of MAGUKs, and Ca2+-activated calmodulin may serve as a general regulator to affect the interactions of MAGUKs and various components of the cytoskeleton

Could peak proteinuria determine whether patient with dengue fever develop dengue hemorrhagic/dengue shock syndrome? - A prospective cohort study

BACKGROUND: Worldwide there is a need to develop simple effective predictors that can distinguish whether a patient will progress from dengue fever (DF) to life threatening dengue hemorrhagic (DHF) or dengue shock syndrome (DSS). We explored whether proteinuria could be used as such a marker. METHODS: We included patients admitted to hospital with suspected dengue fever. Starting at enrollment until discharge, each patient's daily spot urine protein creatinine ratio (UPCR) was measured. We classified those with confirmed dengue infection as DF or DHF (including DSS) based on WHO criteria. Peak and day of onset of proteinuria was compared between both groups. RESULTS: Compared to those with DF, patients with DHF had significantly higher median peak proteinuria levels (0.56 versus 0.08 g/day; p < 0.001). For patients with DHF, the median day of onset of proteinuria was at 6 days of defervescence, with a range of -2 to +3 days after defervescence. There were three patients with DF who did not have proteinuria during their illness; the five remaining patients with DF had a median day of onset of proteinuria of was at 6 days of defervescence with a range of 0 to +28 days. CONCLUSIONS: Peak UPCR could potentially predict DHF in patients with dengue requiring close monitoring and treatment