Functional Diversity and Structural Disorder in the Human Ubiquitination Pathway

The ubiquitin-proteasome system plays a central role in cellular regulation and protein quality control (PQC). The system is built as a pyramid of increasing complexity, with two E1 (ubiquitin activating), few dozen E2 (ubiquitin conjugating) and several hundred E3 (ubiquitin ligase) enzymes. By collecting and analyzing E3 sequences from the KEGG BRITE database and literature, we assembled a coherent dataset of 563 human E3s and analyzed their various physical features. We found an increase in structural disorder of the system with multiple disorder predictors (IUPred - E1: 5.97%, E2: 17.74%, E3: 20.03%). E3s that can bind E2 and substrate simultaneously (single subunit E3, ssE3) have significantly higher disorder (22.98%) than E3s in which E2 binding (multi RING-finger, mRF, 0.62%), scaffolding (6.01%) and substrate binding (adaptor/substrate recognition subunits, 17.33%) functions are separated. In ssE3s, the disorder was localized in the substrate/adaptor binding domains, whereas the E2-binding RING/HECT-domains were structured. To demonstrate the involvement of disorder in E3 function, we applied normal modes and molecular dynamics analyses to show how a disordered and highly flexible linker in human CBL (an E3 that acts as a regulator of several tyrosine kinase-mediated signalling pathways) facilitates long-range conformational changes bringing substrate and E2-binding domains towards each other and thus assisting in ubiquitin transfer. E3s with multiple interaction partners (as evidenced by data in STRING) also possess elevated levels of disorder (hubs, 22.90% vs. non-hubs, 18.36%). Furthermore, a search in PDB uncovered 21 distinct human E3 interactions, in 7 of which the disordered region of E3s undergoes induced folding (or mutual induced folding) in the presence of the partner. In conclusion, our data highlights the primary role of structural disorder in the functions of E3 ligases that manifests itself in the substrate/adaptor binding functions as well as the mechanism of ubiquitin transfer by long-range conformational transitions. © 2013 Bhowmick et al

Common Genetic Denominators for Ca++-Based Skeleton in Metazoa: Role of Osteoclast-Stimulating Factor and of Carbonic Anhydrase in a Calcareous Sponge

Calcium-based matrices serve predominantly as inorganic, hard skeletal systems in Metazoa from calcareous sponges [phylum Porifera; class Calcarea] to proto- and deuterostomian multicellular animals. The calcareous sponges form their skeletal elements, the spicules, from amorphous calcium carbonate (ACC). Treatment of spicules from Sycon raphanus with sodium hypochlorite (NaOCl) results in the disintegration of the ACC in those skeletal elements. Until now a distinct protein/enzyme involved in ACC metabolism could not been identified in those animals. We applied the technique of phage display combinatorial libraries to identify oligopeptides that bind to NaOCl-treated spicules: those oligopeptides allowed us to detect proteins that bind to those spicules. Two molecules have been identified, the (putative) enzyme carbonic anhydrase and the (putative) osteoclast-stimulating factor (OSTF), that are involved in the catabolism of ACC. The complete cDNAs were isolated and the recombinant proteins were prepared to raise antibodies. In turn, immunofluorescence staining of tissue slices and qPCR analyses have been performed. The data show that sponges, cultivated under standard condition (10 mM CaCl2) show low levels of transcripts/proteins for carbonic anhydrase or OSTF, compared to those animals that had been cultivated under Ca2+-depletion condition (1 mM CaCl2). Our data identify with the carbonic anhydrase and the OSTF the first two molecules which remain conserved in cells, potentially involved in Ca-based skeletal dissolution, from sponges (sclerocytes) to human (osteoclast)

Computing internodal conductivities in numerical modeling of two dimensional unsaturated flow on rectangular grid

This paper compares numerical solutions of transient two-dimensional unsaturated flow equation by using different averaging schemes for internodal conductivities. Averaging methods such as arithmetic mean, geometric mean, upstream weighting, and integrated mean are taken into account, as well as a recent approach based on steady-state approximation. The latter method proved the most flexible, producing relatively accurate solutions for both downward and upward flow cases.W artykule porównano rozwiązania numeryczne dwuwymiarowego równania przepływu nienasyconego z użyciem różnych metod uśredniania współczynnika przewodności hydraulicznej między sąsiednimi węzłami. Uwzględniono średnią arytmetyczną, geometryczną, 'pod prąd' oraz całkową, a także niedawno zaproponowany sposób uśredniania oparty na analizie przepływu ustalonego. Ten ostatni sposób okazał się najbardziej uniwersalny, gdyż umożliwiał otrzymanie stosunkowo dokładnych wyników dla przepływu wody w strefie nienasyconej zarówno w kierunku w dół jak i w górę

Simulation of water flow in double-porosity soils on unstructured grids with the finite-volume method

Double-porosity soils consist of two interacting porous systems corresponding to weakly conductive aggregates and highly conductive inter-aggregate regions. The flow of water in such media can be described with a two-scale model obtained by homogenization. The model consists of a single macroscopic equation for the flow in the highly conductive porous system coupled with a number of micro-scale equations for the flow in the weakly conductive aggregates. In this paper we present a numerical algorithm to solve the resulting system of equations for the case of macroscopically two-dimensional flow. It is based on the finite volume approach for unstructured grid of triangular cells. Special attention is paid to the coupling of the micro- and macro-scale equations. An exemplary calculation is presented, concerning infiltration and redistribution of water in a hill-slope of double-porosity structure with cubic aggregates

Unstructured finite-volume meshes for two-dimensional flow in variably saturated porous media

This paper presents a numerical algorithm for solving the equation describing variably saturated flow in porous media. The algorithm is based on a control volume finite element approach and can be applied to two-dimensional unstructured meshes consisting of triangular elements. Two methods of defining the dual control volume grid are discussed. We also demonstrate that the method of calculating the average permeability at the control volume face significantly influences numerical results

Computing Internodal Conductivities in Numerical Modeling of two Dimensional Unsaturated Flow on Rectangular Grid

W artykule porównano rozwiazania numeryczne dwuwymiarowego równania przepływu nienasyconego z uzyciem róznych metod usredniania współczynnika przewodnosci hydraulicznej miedzy sasiednimi wezłami. Uwzgledniono srednia arytmetyczna, geometryczna, “pod prad” oraz całkowa, a takze niedawno zaproponowany sposób usredniania oparty na analizie przepływu ustalonego. Ten ostatni sposób okazał sie najbardziej uniwersalny, gdyz umozliwiał otrzymanie stosunkowo dokładnych wyników dla przepływu wody w strefie nienasyconej zarówno w kierunku w dół jak i w góre