Coupling of folding and binding in the PTB domain of the signaling protein Shc

The notion that certain proteins lack intrinsic globular structure under physiological conditions and that the attainment of fully folded structure only occurs upon the binding of target molecules has been recently gaining popularity. We report here the solution structure of the PTB domain of the signaling protein Shc in the free form. Comparison of this structure with that of the complex form, obtained previously with a phosphopeptide ligand, reveals that the Shc PTB domain is structurally disordered in the free form, particularly around the regions constituting the peptide binding pocket. The binding of the ligand appears to reorganize this pocket through local folding events triggering a conformational switch between the free and the complex forms.     

Solution structure of the MAPK phosphatase PAC-1 catalytic domain. Insights into substrate-induced enzymatic activation of MKP

Inactivation of mitogen-activated protein kinases (MAPKs) by MAPK phosphatases (MKPs) is accomplished via substrate-induced activation of the latter enzymes; however, the structural basis for the underlying mechanism remains elusive. Here, we report the three-dimensional solution structure of the C-terminal phosphatase domain of the prototypical MKP PAC-1, determined when bound to phosphate. Structural and biochemical analyses reveal unique active site geometry of the enzyme important for binding to phosphorylated threonine and tyrosine of MAPK ERK2. Our study further demonstrates that the dynamic interaction between the N-terminal kinase binding domain and the C-terminal phosphatase domain of an MKP is directly coupled to MAPK-induced conformational change of the phosphatase active site, which is essential for eliciting its full enzymatic activity.     

The Nef protein of HIV-1 induces loss of cell surface costimulatory molecules CD80 and CD86 in APCs

The Nef protein of HIV-1 is essential for its pathogenicity and is known to down-regulate MHC expression on infected cell surfaces. We now show that Nef also redistributes the costimulatory molecules CD80 and CD86 away from the cell surface in the human monocytic U937 cell line as well as in mouse macrophages and dendritic cells. Furthermore, HIV-1-infected U937 cells and human blood-derived macrophages show a similar loss of cell surface CD80 and CD86. Nef colocalizes with MHC class I (MHCI), CD80, and CD86 in intracellular compartments, and binds to both mouse and human CD80 and CD86. Some Nef mutants defective in MHCI down-modulation, including one from a clinical isolate, remain capable of down-modulating CD80 and CD86. Nef-mediated loss of surface CD80/CD86 is functionally significant, because it leads to compromised activation of naive T cells. This novel immunomodulatory role of Nef may be of potential importance in explaining the correlations of macrophage-tropism and Nef with HIV-1 pathogenicity and immune evasion.     

Age-related peculiarities of change in content of free radical oxidation products in muscle during stress

The age-dependent peculiarities of stimulation of free radical processes in subcellular fractions of skeletal muscle of rats subjected to long-term immobilization stress were studied in order to improve knowledge about changes of muscular tissue during ontogenesis. It is found that adult animals do not show accumulation of proteins carbonyls, TBA-reactive substances, and Schiff bases in subcellular fractions of the thigh muscle when immobUized. Long-term immobilization causes apparent manifestation of oxidative stress only in mitochondrial fraction in pubertal rats. Mitochondrial oxidative stress defense systems are sufficiently effective, however, direction of pathways of free radical oxidation carbonyl products catabolism alters in the cytoplasm of myocytes in old rats under long-term immobilization conditions.