Analysis of binding properties and specificity through identification of the interface forming residues (IFR) for serine proteases <Emphasis Type="Italic">in silico</Emphasis> docked to different inhibitors

Background  

Enzymes belonging to the same super family of proteins in general operate on variety of substrates and are inhibited by wide selection of inhibitors. In this work our main objective was to expand the scope of studies that consider only the catalytic and binding pocket amino acids while analyzing enzyme specificity and instead, include a wider category which we have named the Interface Forming Residues (IFR). We were motivated to identify those amino acids with decreased accessibility to solvent after docking of different types of inhibitors to sub classes of serine proteases and then create a table (matrix) of all amino acid positions at the interface as well as their respective occupancies. Our goal is to establish a platform for analysis of the relationship between IFR characteristics and binding properties/specificity for bi-molecular complexes.     

Subcellular distribution of clathrin in cultured hypothalamic neurons.

The subcellular distribution of clathrin has been examined in developing hypothalamic neurons cultured in a chemically defined medium up to synapse formation (12-13 days in vitro) and exposed, or not, to a depolarizing concentration of KCl (60 mM for 3 min) followed, or not, by a return to control KCl concentration (3 mM KCl for 3 min). Previous studies have shown that such treatments induce in synaptic boutons a rapid vesicle depletion followed by massive restoration. Using an enzyme immunoassay, we have compared the relative proportion of assembled and unassembled pools of clathrin as a function of exposure to depolarizing or repolarizing concentrations of KCl. In parallel we have localized clathrin at the electron microscopic level using immunoperoxidase. Clathrin concentration in culture is lower (0.36 vs 0.75%) and the proportion of unassembled clathrin is much higher than in the adult brain (82 vs 14%). These proportions were not affected by depolarizing or repolarizing treatments. Morphologically clathrin was exclusively detected in two neuron compartments: perikarya and synaptic boutons. In perikarya clathrin was localized as a thick coat on plasma membrane coated pits and in the Golgi zone on coated buds and vesicles, presumably located in a trans compartment. In synaptic boutons clathrin immunoreaction was found as an irregular thin rim around synaptic vesicles, whatever the polarization state of the cells, but coated vesicles were extremely rare. Taken together these findings raise the problem of the functional meaning and localization of the large unassembled pool of clathrin in such neurons and question its role in vesicular traffic in synaptic boutons.     

Golgi localisation of GMAP210 requires two distinct cis-membrane binding mechanisms

Background  

The Golgi apparatus in mammals appears as a ribbon made up of interconnected stacks of flattened cisternae that is positioned close to the centrosome in a microtubule-dependent manner. How this organisation is achieved and retained is not well understood. GMAP210 is a long coiled-coil cis-Golgi associated protein that plays a role in maintaining Golgi ribbon integrity and position and contributes to the formation of the primary cilium. An amphipathic alpha-helix able to bind liposomes in vitro has been recently identified at the first 38 amino acids of the protein (amphipathic lipid-packing sensor motif), and an ARF1-binding domain (Grip-related Arf-binding domain) was found at the C-terminus. To which type of membranes these two GMAP210 regions bind in vivo and how this contributes to GMAP210 localisation and function remains to be investigated.     

Genetic structure of three Naikpod subpopulations of Andhra Pradesh, India

The genetic structure of three subpopulations of the Naikpod tribe of Andhra Pradesh, India, was examined by studying three blood group, six red cell enzyme, and five protein systems and phenylthiocarbamide taste sensitivity. The gene frequency data of 15 loci are compared among the subpopulations as well as with those reported for other population groups from India. The analysis of gene diversity revealed that the gene differentiation among the subpopulations relative to total population is only 0.02, indicating that the genetic differentiation between subpopulations is very small compared with that within them. This is corroborated by the small genetic distances found among them. The effect of differentiation of microgeographical and breeding isolation on gene diversity and genetic differentiation among the three subpopulations is apparently low.     

Gc subtyping in south Indian tribal and caste populations

Seven tribal (Konda Kammara - 2 samples; Koya Dora - 3 samples; Lambadi) and caste (Madiga) populations from Andhra Pradesh (South India) have been analyzed for the distribution of Gc subtypes. The observed heterogeneity in the distribution of Gc1F, Gc1S and Gc2 alleles was found to be statistically significant. Comparisons are made with North Indian populations as well as with those of other racial affiliation. The anthropological impact of the Gc subtype polymorphism is discussed.