Targeting Domain

Probably the smallest sequence known to be responsible for receptor recognition is the RGD-tripeptide, initially discovered in fibronectin [143]. However, the specificity of the interaction with different integrins, the counter receptors of RGD sequences on the cell surface, is established by the flanking sequences of the RGD motif and the conformation of the tripeptide. In other words, the presentation of the RGD sequence is important for specific recognition by individual integrins. 

The use of phage-display techniques has identified peptide ligands with specific affinity to cell surface receptors or specific tissues (see Table 11.2). Such peptide homing devices can be 

---

FAT Focal adhesion targeting domain Binding to focal adhesions complexes... 

Munro, S., and Nichols, B. (1999). The grip domain—a novel Golgi-targeting domain found in several coiled-coil proteins. Gun. Biol. 9, 377—380. 

The functions of these fusion proteins and their precise roles in the induction of cancer are currently the subject of intense investigation (Collins et al., 2006 Huntly et al., 2004 Kindle et al., 2005). In general, it appears that since the proteins are fused in-frame, their catalytic domains continue to function, but these activities are mis-directed by the targeting domains of their fusion partners. The result is aberrant acetylation of chromatin and non-chromatin proteins, as well as sequestration of key nuclear regulators. These mechanisms are discussed in-depth in Chapter 8 of this book. Furthermore, two recent studies have demonstrated that wild-type MOZ is essential for development of hematopoietic stem cells (Katsumoto et al., 2006 Thomas et al., 2006). This suggests that AML may arise not only due to the aberrant activity of the fusion proteins, but also due to the loss of their original function. 

A subsequent study in 2002 of 27 families with a condition known as multiminicore disease (MmD) also linked mutations in SEPNl to disease pathology. Multiple mutations were identified in exons 1, 5, 7, 8, 10, and 11, and the authors also mentioned that this region (RSMD) had been previously linked to MmD. Minicores are lesions by histochemistry of mitochondrial depletion within muscle tissue. The first biochemical study of selenoprotein N aimed to identify the protein localization by immunohistochemistry and found that the primary protein product of several identified mRNAs (splice variants) was a 70 kDa protein present in the endoplasmic reticulum. Two potential ER targeting domains were shown to be present and the peptide expressed from the first exon was shown to be required for localization into the ER. This study also revealed that selenoprotein N was an integral membrane protein that is N-glycosylated. Expression analysis showed pronounced levels in embryonic tissue with a reduction after development and differentiation. 

Another level of specificity of DUBs is with respect to the substrates that are deubiquitinated. Although not much is known about how DUBs choose their targets, domains outside the catalytic core might play a role in determining the substrate to which a given DUB binds. For example, a CAP-Gly domain (a conserved glycine-rich domain found in some cytoskeleton-associated proteins (CAPs)) of CYLD functions in its interaction with NF-kB essential modulator (NEMO), a putative CYLD substrate. 

In terms of targeting, membrane translocation domains lack specificity for particnlar cells or tissues. Therefore, these domains shonld be combined with targeting domains snch as those discussed in the previous section. In snch a constrnct, the targeting domain wiU ensnre a rapid accumulation at the surface of a specific cell type and the translocation domain will facilitate entry into the cytosol of the target cells. 

The sequence of bovine ADPH was determined from its cDNA (Gen-bank accession number AJ011680) the bovine protein is 87 and 80% identical to human and mouse ADPH, respectively (Nielsen et al., 1999). Surprisingly, ADPH does not have an extensive contiguous sequence of hydrophobic amino acids. About half of the 50 N-terminal residues are nonpolar. McMa-naman et al. (2003) found that the lipid-targeting domain of ADPH is in a presumptive u-helical region between residues 189 and 205 of the mouse protein (Figure 4.9). 

The dependent measure was the time to read the last (metaphorical) sentence. To ensure comparability, this final test sentence was always the same the initial setting passage was varied between conditions. In all cases the same meaning in the target domain was conveyed in the two... 

Plants also produce structurally related enzymes (chitinases) that catalyse the hydrolysis of chitin (Table 12.2) and hence damage chitin-based insect integuments. Class I chitinases are basic enzymes with an jV-terminal hevein-related CBD and vacuole-targeting C-terminal signals whereas Class II enzymes are acidic proteins lacking these CBD and vacuole-targeting domains. Class IV chitinases are variously basic and acidic extracellular proteins with... 

Fig. 4. Multiple sequence alignment of Atx 1 and N-terminal domains of the P-type ATPases using the CLUSTAL method (Higgins and Sharp, 1989). Sequence numbering corresponds to that of the yeast proteins. (Top) The copper chaperones are shown and (bottom) the N-terminal domains of the target P-type ATPases are shown. The sulfur-containing components of the MXCXXC motifs are boxed in black between residues 10 and 20. Residues thought to be important in the electrostatic recognition between chaperones and target domains are boxed in black and labeled with an asterisk (see text).

Sequence References for Atxl-like Copper Chaperones and Their Target Domains... 

Fig. 5. Three-dimensional structures of the (iaPPaP proteins oftheAtxl-like family. MXCXXC motif residues are hoxed. The Protein Data Bank (pdh) code for each structure is in parentheses, (a) NMR structure of Shigella flexneri Hg(II)MerP (Steele and Opella, 1997). (h) X-ray structure oiSaccharamyces cerevisiae Hg(II)Atxl. K24, K28, K59, and K62, side chains important in the recognition of the Ccc2a target domain, are shown outside of the hox (see text) (Rosenzweig et al., 1999). (c) X-ray structure of human Cu(I)Hahl. R21, K25, K56, and K57, side chains important in the recognition of the fourth N-terminal domain of the Menkes protein, are shown outside of the box (Wernimont et al., 2000). (d) NMR structure oi Enterococcus hirae apoCopZ (Wimmer et al., 1999). (e) NMR structure of human Ag(l)Mnk4, the fourth domain of the...

Based on the wealth of both in vivo and in vitro biochemical experiments described in Section II, A, a diffusion-driven or bucket brigade mechanism of metal ion transfer between MXCXXC-motifs on the copper chaperone proteins and their target domains was predicted (Pufahl et al.. 

Fig. 3.8 The protein tyrosine phosphatases (PTPs) with the C(X)gR motif are divided into tyrosine-specific PTPs the VHl-like dual specificity, serine/threonine- and tyrosine-specific phosphatases the CDC25 phosphatase and the low molecular weight (LMW) phosphatases. The low molecular weight phosphatases are acid phosphatases without distinct regulatory or targeting domains. Their function is not known. The tyosiiie-speoifio phosphatases are further subdivided into receptor-like and non-reoeptor-like phosphatases. (This scheme is reproduced with permission of the authors and Trends Biochem. Sci. from ref. 76.)...

Fig. 3.1. Posttranslational modifications of Ras superfamily small GTPases in which both prenylation and C-terminal phosphorylation have been demonstrated. The sequences of the hypervariable (membrane targeting) domain (HVD) of each such GTPase are shown. Known phosphorylation sites are bolded and shown in red numbers are amino acid residues. Basic residues (with a positive charge) are shown in blue. Responsible kinases are indicated. S, serine T, threonine Y, tyrosine.

---