An EVH1/WH1 domain as a key actor in TGFβ signalling

AbstractEVH1 (enabled VASP (vasodilator-stimulated protein) homology 1)/WH1 (WASP (Wiskott–Aldrich syndrome protein) homology 1) domains, present in Ena VASP and WASP, are protein interaction modules specialised in binding proline-rich ligands. An EVH1/WH1 domain is here identified in the recently cloned SMIF protein, a key protein in transforming growth factor-β (TGFβ) signalling which was not yet related to defined domains. The SMIF EVH1/WH1 domain interacts with the proline-rich Smad4 activation domain, leading to translocation of so-formed complex to the nucleus where SMIF possesses strong intrinsic TGFβ-inducible transcriptional activity. This finding highlights the pivotal role that the EVH1/WH1 family of domains play in multiple eukaryotic signal transduction pathways

Trigger factor, one of the Escherichia coli chaperone proteins, is an original member of the FKBP family

AbstractThe trigger factor of Escherichia coli is known as a chaperone protein which forms soluble complexes with the precursor to outer membrane protein A and assists in the maintenance of translocation competence. Sequence analysis shows that trigger factor contains a domain belonging to the FK506-binding protein (FKBP) family and possessing all the amino acids necessary for FK506 binding and peptidyl-prolyl cis-trans isomerase (Ppiase) activity. Consequently, this protein could be directly involved in the unfolding/folding processes occurring during translocation across the E. coli plasma membrane and, more generally, in facilitating protein folding. The central position of the FKBP domain within the trigger factor sequence as well as several original features of the loops surrounding the FK506-binding pocket are not found in any other FKBPs, making it undetectable by the Fkbp-Ppiase signature patterns

Investigation of MIG welding to the corrosion behaviour of carbon steel

A welding process was performed on the low carbon steel based on different welding parameters in this experiment. The main objective of this study was to investigate the effect of welding voltages (19 V, 20 V, 21 V, and 22 V) and wire feed rates (90 ipm, 100 ipm, 110 ipm, and 120 ipm) to the corrosion behaviour of welded carbon steel in synthetic seawater environment (3.5 wt% NaCl). Besides that, the microstructure changes in the weldments of maximum and minimum voltages were analysed and compared with the as-received sample before welding process. On the other hand, the penetration level of the weldments was analysed due to the different welding parameter. The welding method in this experiment was metal inert gas (MIG) with filler metal (ER 70S-6 with 1.2 mm diameter). Based on the results obtained, the corrosion rate decreased when the welding voltage was increased from 19 V to 22 V or when the wire feed rate was increased from 90 ipm to 120 ipm until the full penetration level of weldment occurred. In conclusion, the lowest corrosion rate occurred at highest welding voltage and highest wire feed rate when full penetration of weldment occurred. The corrosion product that formed on the surface of the sample was iron oxides and pitting was found on the surface of the exposed area after the corrosion test

Characterization of Non-Trivial Neighborhood Fold Constraints from Protein Sequences using Generalized Topohydrophobicity

Prediction of key features of protein structures, such as secondary structure, solvent accessibility and number of contacts between residues, provides useful structural constraints for comparative modeling, fold recognition, ab-initio fold prediction and detection of remote relationships. In this study, we aim at characterizing the number of non-trivial close neighbors, or long-range contacts of a residue, as a function of its “topohydrophobic” index deduced from multiple sequence alignments and of the secondary structure in which it is embedded. The “topohydrophobic” index is calculated using a two-class distribution of amino acids, based on their mean atom depths. From a large set of structural alignments processed from the FSSP database, we selected 1485 structural sub-families including at least 8 members, with accurate alignments and limited redundancy. We show that residues within helices, even when deeply buried, have few non-trivial neighbors (0–2), whereas β-strand residues clearly exhibit a multimodal behavior, dominated by the local geometry of the tetrahedron (3 non-trivial close neighbors associated with one tetrahedron; 6 with two tetrahedra). This observed behavior allows the distinction, from sequence profiles, between edge and central β-strands within β-sheets. Useful topological constraints on the immediate neighborhood of an amino acid, but also on its correlated solvent accessibility, can thus be derived using this approach, from the simple knowledge of multiple sequence alignments

Identification of a new expanding family of genes characterized by atypical LRR domains. Localization of a cluster preferentially expressed in oocyte

AbstractIn the present work, we have used the in silico subtraction methodology to identify novel oocyte-specific genes in the mouse. By this way, we have identified in silico a new family of genes composed of more than 80 members. Sequence analysis showed that these genes belong to the superfamily of leucine-rich repeat (LRR) proteins. However, LRRs of this family display some variability in length and in amino acids composition within the β-strands region, as more leucine residues are substituted by other hydrophobic amino acids as compared to canonical LRRs. Interestingly, for nine of these genes, the ESTs were represented almost exclusively in mouse egg libraries. Three of them were selected for experimental study. By RT-PCR and in situ hybridization, we confirmed their specific expression in the mouse oocyte from primary to preovulatory follicles. These three genes are localized in a cluster on mouse chromosome 4, in the vicinity of another recently discovered oocyte specific gene called oogenesin, that we also found to belong to the same family. We thus re-named this latter gene ‘oogenesin-1’, and the three genes identified here were named oogenesin-2, -3 and -4

The Metallo-β-Lactamase/β-CASP Domain of Artemis Constitutes the Catalytic Core for V(D)J Recombination

The V(D)J recombination/DNA repair factor Artemis belongs to the metallo-β-lactamase (β-Lact) superfamily of enzymes. Three regions can be defined within the Artemis protein sequence: (a) the β-Lact homology domain, to which is appended (b) the β-CASP region, specific of members of the β-Lact superfamily acting on nucleic acids, and (c) the COOH-terminal domain. Using in vitro mutagenesis, here we show that the association of the β-Lact and the β-CASP regions suffices for in vivo V(D)J recombination of chromosome-integrated substrates. Single amino acid mutants point to critical catalytic residues for V(D)J recombination activity. The results presented here define the β-Lact/β-CASP domain of Artemis as the minimal core catalytic domain needed for V(D)J recombination and suggest that Artemis uses one or two Zn(II) ions to exert its catalytic activity, like bacterial class B β-Lact enzymes hydrolyzing β-lactam compounds

Prediction of the general transcription factors associated with RNA polymerase II in Plasmodium falciparum: conserved features and differences relative to other eukaryotes

BACKGROUND: To date, only a few transcription factors have been identified in the genome of the parasite Plasmodium falciparum, the causative agent of malaria. Moreover, no detailed molecular analysis of its basal transcription machinery, which is otherwise well-conserved in the crown group of eukaryotes, has yet been reported. In this study, we have used a combination of sensitive sequence analysis methods to predict the existence of several parasite encoded general transcription factors associated with RNA polymerase II. RESULTS: Several orthologs of general transcription factors associated with RNA polymerase II can be predicted among the hypothetical proteins of the P. falciparum genome using the two-dimensional Hydrophobic Cluster Analysis (HCA) together with profile-based search methods (PSI-BLAST). These predicted orthologous genes encoding putative transcription factors include the large subunit of TFIIA and two candidates for its small subunit, the TFIIE β-subunit, which would associate with the previously known TFIIE α-subunit, the TFIIF β-subunit, as well as the p62/TFB1 subunit of the TFIIH core. Within TFIID, the putative orthologs of TAF1, TAF2, TAF7 and TAF10 were also predicted. However, no candidates for TAFs with classical histone fold domain (HFD) were found, suggesting an unusual architecture of TFIID complex of RNA polymerase II in the parasite. CONCLUSION: Taken together, these results suggest that more general transcription factors may be present in the P. falciparum proteome than initially thought. The prediction of these orthologous general transcription factors opens the way for further studies dealing with transcriptional regulation in P. falciparum. These alternative and sensitive sequence analysis methods can help to identify candidates for other transcriptional regulatory factors in P. falciparum. They will also facilitate the prediction of biological functions for several orphan proteins from other apicomplexan parasites such as Toxoplasma gondii, Cryptosporidium parvum and Eimeria

A generalized analysis of hydrophobic and loop clusters within globular protein sequences

BACKGROUND: Hydrophobic Cluster Analysis (HCA) is an efficient way to compare highly divergent sequences through the implicit secondary structure information directly derived from hydrophobic clusters. However, its efficiency and application are currently limited by the need of user expertise. In order to help the analysis of HCA plots, we report here the structural preferences of hydrophobic cluster species, which are frequently encountered in globular domains of proteins. These species are characterized only by their hydrophobic/non-hydrophobic dichotomy. This analysis has been extended to loop-forming clusters, using an appropriate loop alphabet. RESULTS: The structural behavior of hydrophobic cluster species, which are typical of protein globular domains, was investigated within banks of experimental structures, considered at different levels of sequence redundancy. The 294 more frequent hydrophobic cluster species were analyzed with regard to their association with the different secondary structures (frequencies of association with secondary structures and secondary structure propensities). Hydrophobic cluster species are predominantly associated with regular secondary structures, and a large part (60 %) reveals preferences for α-helices or β-strands. Moreover, the analysis of the hydrophobic cluster amino acid composition generally allows for finer prediction of the regular secondary structure associated with the considered cluster within a cluster species. We also investigated the behavior of loop forming clusters, using a "PGDNS" alphabet. These loop clusters do not overlap with hydrophobic clusters and are highly associated with coils. Finally, the structural information contained in the hydrophobic structural words, as deduced from experimental structures, was compared to the PSI-PRED predictions, revealing that β-strands and especially α-helices are generally over-predicted within the limits of typical β and α hydrophobic clusters. CONCLUSION: The dictionary of hydrophobic clusters described here can help the HCA user to interpret and compare the HCA plots of globular protein sequences, as well as provides an original fundamental insight into the structural bricks of protein folds. Moreover, the novel loop cluster analysis brings additional information for secondary structure prediction on the whole sequence through a generalized cluster analysis (GCA), and not only on regular secondary structures. Such information lays the foundations for developing a new and original tool for secondary structure prediction