Protein domain orientation

Altogether, the identification of the coordinating residues in the endogenous hDAT Zn2+ binding site followed by the engineering artificial sites have defined an important series of structural constraints in this transporter. This includes not only a series of proximity relationships in the tertiary structure, but also secondary structure relationships. The data also provided information about the orientation of TM7 relative to TM8. A model of the TM7/8 microdomain that incorporates all these structural constraints is shown in Fig. 4 (36). The model is an important example of how structural inferences derived from a series of Zn2+ binding sites can provide sufficient information for at least an initial structural mapping of a selected protein domain. 

Eukaryotic homology, gene-orientated clusters of transcript sequences, conserved protein domain, markers and mapping, population study (PopSet), expression and molecular abundance profiles (GEO), and GEO data sets and cancer chromosomes. 

Sander and Schneider performed an empirical determination of homology thresholds by studying thousands of sequence alignments within the PDB database [19]. One has to keep in mind that perfect sequence similarity not always implies perfect structural agreement a protein crystal structure may vary, typically in loop regions or in domain orientation, as a result of different substrate or cofactor interaction, complex formation or crystal contacts. The curve in Figure 4, however, provides a reliable rule of thumb for those who wonder if their enzyme can be modeled with any degree of confidence. 

Contributions by R. Joseph and P. Arya as well as M. A. Koch and H. Waldmann focus on synthetic aspects towards lead structures originating from natural product-derived scaffolds. R. Joseph and P. Arya refer to two complementary approaches, the synthetic access to focussed libraries around bioactive natural product cores, and diversity-oriented synthesis aiming at 3D scaffold diversity for hit generation, respectively. On the other hand, M. A. Koch and H. Waldmann emphasise the correlation of natural product-based library concepts with structural features of targeted protein domains, thus strengthening the privileged structure concept from a bioorganic viewpoint. 

Many fundamental biological processes appear to depend on unique properties of inner hydrophilic domains of the membrane proteins in which ions or water molecules diffuse along the directional pathways. In the membrane protein systems simple inner functional moieties (i.e. carbonyl, hydroxyl, amide, etc.) are pointing toward the protein core, surrounded by the outer scaffolding protein wall orienting the transport direction. Bilayer or nanotube artificial membranes were developed with the hope of mimicking the natural ion-channels, which can directly benefit the fields of separations, sensors or storage-delivery devices. 

Hwang, P. M., Skrynnikov, N. R., and Kay, L. E. (2001). Domain orientation in beta-cyclodextrin-loaded maltose binding protein Diffusion anisotropy measurements confirm the results of a dipolar coupling study. J. Biomol. NMR 20, 83-88. 

The three-dimensional structure of the PapD periplasmic chaperone that forms transient complexes with pilus subunit proteins has been solved by Holmgren and Branden (1989). PapD consists of two globular domains oriented in the shape of a boomerang (Fig. 2). Each domain is a /3-barrel structure formed by two antiparallel /8-pleated sheets that have a topology similar to an immunoglobulin fold. The relationship between PapD and other immunoglobulin-like proteins is discussed in Section IV,C. 

Figure 16 Structure calculation purely based on NOE and dihedral angle restrictions often fails to determine proper domain orientations. By incorporating the long-range orientation parameters that RDCs provide, it is possible to overcome this problem and to define relative orientation of the two domains of the ribosomal LI 1 protein.

The same method has been applied to measure the Eli domain orientation when the protein is in complex with its RNA parmer or both RNA and thiostrepton antibiotic. The additional RDCs revealed a rearrangement of the N-terminal domain of Ell placing it closer to the RNA after binding of thiostrepton. HADDOCK has been used to calculate a model of the ternary stmcture of the Ell protein in complex with RNA and antibiotic. Based on the orientational data, the dynamics and the docking model, it seems that thiostrepton locks the domain conformation of Ell in a rigid (inhibitory) state. The antibiotic thiostrepton interferes with the interaction of elongation factors to this Ell-RNA region, which has a dramatic effect on the level of protein synthesis by the ribosome. 

In many cases, small molecules are found for a TF of interest through assays developed with known protein partners. Structures of the interacting protein domains also have been invaluable for drug discovery. For several important TFs, however, the specific cellular partners with which they interact are unknown. In this case, using small molecule screens on solid supports is an extremely useful tool for dmg discovery. In one such screen, diversity-oriented synthesis (DOS (129) was used to identify a small molecule binder of Hap3p, a yeast protein... 

A graphical approach was also used by Millet and Pons to analyse anisotropy of rotational diffusion in proteins. The values of Z)j and DJD compatible with R IRi ratios are presented as a contour plot. The intersection of the contour plots for different residues provides the values of anisotropy parameters compatible with experimental data. The obtained parameters can be used as starting values for further optimisation. The method is apphcable to axially symmetric rotation. A combination of approximate and exact methods was used by Ghose et al. to reduce the computational time of the determination of rotational diffusion tensor from NMR relaxation data. The initial values of the tensor components and its orientation are evaluated from the approximate solution, which substantially reduces the search space during the exact calculations. The method was applied for the estimation of relative domain orientation of a dual domain protein. 

Xie, J., Bogdanov, M., Heacock, R, Dowhan, W. 2008. To flip or not to flip protein-lipid chaige interactions are determinants of transmembrane domain orientation. J. Biol. Chem. Submitted. 

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