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In this report, an important advance in understanding the complexities of protein splicing is made possible by the manipulation of the trans -splicing intein involved in the maturation of the Synechocystis sp. Volume 40, Issue 2, 1 MayPages e10—e Zinc inhibition, together with the reversibility of EDTA, could provide a novel approach to modulate protein splicing and peptide bond cleavage. Furthermore, the Ssp DnaE intein possesses neither a His residue at the penultimate position nor a Glu residue at the equivalent position Glu 80 in block B, which are involved in the coordination of zinc in the x-ray structure of the Sce VMA intein Service delivery systems recommendations include increased social support systems, colocated and integrated services, and training of case managers to motivate clients to seek mental health and substance abuse treatment.
It is possible that chelation of a zinc atom by Cys may exclude the binding of DTT, thereby inhibiting its nucleophilic attack on the thioester bond. I have read the information. Remarkably, a zinc atom was found at the catalytic center of this splicing-deficient precursor protein. The effect of zinc ion on cleavage at the N-terminal splice junction, the initial step in the splicing pathway, was examined in the presence of 5 m m DTT.
Examination of the interaction between the N- and C-terminal intein halves would offer important insight into the tertiary folding that brings all the reacting groups in close proximity. Immunoblot analysis of protein trans- splicing activity of various intein mutants.
Removal of more than 3 residues, following the translation initiation methionine of I Cresulted in complete inhibition of growth of the host cells, suggesting that an active ALS was not produced. Search Enter search terms: Furthermore, site-directed mutagenesis was performed to study the effect of amino acid substitutions in the highly conserved intein blocks B in I N and F in I C on the production of an active ALS.
N and C termini of the intein are labeled N and Crespectively. The C-terminal 23 residues of I N appear not to be required for either association or splicing, probably because they constitute a flexible linker region Fig. T7 gene1 contains a mutated ALS gene.
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We demonstrated the correlation between the splicing or association functions of the 42051 intein and the reconstitution of Escherichia coli acetolactate synthase isoform II ALS. Services Email this article to a friend Alert me when this article is cited Alert me if a correction is posted Alert me when eletters are published Similar articles in this journal Similar articles in Fs of Science Similar articles in PubMed Download to citation manager Request Permissions.
Protein expression was induced with 0. Further understanding of the mode of zinc binding relies on a precise crystal structure of the Ssp DnaE intein.
Proposed mechanism for protein trans- splicing and its inhibition by zinc involving the Ssp DnaE intein. Therefore, the methods demonstrated here should open up new avenues in the application of self-splicing inteins to express and modify proteins.
In this study, we took advantage of the unique property of the trans -splicing Ssp DnaE intein to explore the effect of zinc on splicing activity. Its innovative formula seizes and removes the film produced by limescale and acid rain residues, thus prolonging the effect of the previously applied vs layers.
Most research on this population accessing specialized services 2401 been described from the clients’ perspectives. These data indicated that the inhibitory effect of zinc ion on trans -splicing is not simply due to the presence of a divalent cation, but is specific to zinc. In contrast, growth of the host cells expressing a mutated I C carrying 1 or 2 amino acid deletion indicated the production of a functional ALS.
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One of the fusions carries a wild-type intein sequence, and the other contains a mutated intein segment, as indicated above each lane.
Already have an account? I agree with the transfer of my data to third parties for their free marketing activities. Co-expression of both ALS-intein fusion genes possessing the wild-type intein sequence rescued ER host cells from growth inhibition by valine. The work presented also demonstrates that the presence of zinc ion blocks cleavage of the peptide bond at the N-terminal splice site Fig.
The in vivo functional screen along with an in vitro assay helped to dissect the regions of the intein essential for splicing and interaction between the two intein halves. Aggiungi alla lista dei desideri.