Browsing by keyword "Electrostatics"
Now showing items 1-2 of 2
-
Crystal structure of the catalytic domain of protein-tyrosine phosphatase SHP-1The crystal structures of the protein-tyrosine phosphatase SHP-1 catalytic domain and the complex it forms with the substrate analogue tungstate have been determined and refined to crystallographic R values of 0.209 at 2.5 A resolution and 0.207 at 2.8 A resolution, respectively. Despite low sequence similarity, the catalytic domain of SHP-1 shows high similarity in secondary and tertiary structures with other protein-tyrosine phosphatases (PTPs). In contrast to the conformational changes observed in the crystal structures of PTP1B and Yersinia PTP, the WPD loop (Trp419-Pro428) in the catalytic domain of SHP-1 moves away from the substrate binding pocket after binding the tungstate ion. Sequence alignment and structural analysis suggest that the residues in the WPD loop, especially the amino acid following Asp421, are critical for the movement of WPD loop on binding substrates and the specific activity of protein-tyrosine phosphatases. Our mutagenesis and kinetic measurements have supported this hypothesis.
-
Transmembrane protein insertion orientation in yeast depends on the charge difference across transmembrane segments, their total hydrophobicity, and its distributionThe determinants of transmembrane protein insertion orientation at the endoplasmic reticulum have been investigated in Saccharomyces cerevisiae using variants of a Type III (naturally exofacial N terminus (Nexo)) transmembrane fusion protein derived from the N terminus of Ste2p, the alpha-factor receptor. Small positive and negative charges adjacent to the transmembrane segment had equal and opposite effects on orientation, and this effect was independent of N- or C-terminal location, consistent with a purely electrostatic interaction with response mechanisms. A 3:1 bias toward Nexo insertion, observed in the absence of a charge difference, was shown to reflect the Nexo bias conferred by longer transmembrane segments. Orientation correlated best with total hydrophobicity rather than length, but it was also strongly affected by the distribution of hydrophobicity within the transmembrane segment. The most hydrophobic terminus was preferentially translocated. Insertion orientation thus depends on integration of responses to at least three parameters: charge difference across a transmembrane segment, its total hydrophobicity, and its hydrophobicity gradient. Relative signal strengths were estimated, and consequences for topology prediction are discussed. Responses to transmembrane sequence may depend on protein-translocon interactions, but responses to charge difference may be mediated by the electrostatic field provided by anionic phospholipids.