Protein attachment

SNAPs is an acronym for soluble NSF attachment proteins. They were originally discovered as cofactors for NSF that mediate the membrane binding of NSF in in vitro transport assays. Several isoforms of SNAPs exist in mammalian cells. SNAPs are also highly conserved proteins. Crystallographic studies indicated that the proteins form a very stiff and twisted sheet that is formed by a series of antiparallel and tightly packed helices connected by short loops. 

Stress fibers are parallel bundles of actin filaments that develop in the cytoplasm of fibroblasts from the cortical actin network in response to mechanical tension. These often bind to the plasma membrane at focal contacts and, through transmembrane linker glycoproteins, to the extracellular matrix. Thus, actin filaments of stress fibers indirectly Join to the inner face of the plasma membrane through molecular assemblies of attachment proteins, which include an actin-capping protein, a-actinin, vinculin, and talin (Small, 1988). 

Other viral targets Attachment proteins Fusion proteins Disassembly/Uncoating... 

Figure 48-5. Schematic representation of fibronectin interacting with an integrin fibronectin receptor situated in the exterior of the plasma membrane of a cell of the ECM and of various attachment proteins interacting indirectly or directly with an actin microfilament in the cytosol. For simplicity, the attachment proteins are represented as a complex.

Matrix Attached protein or affinity ligand, or spacer and affinity ligand Ref. 

Hutchinson, F.J., Francis, S.E., Lyle, I.G., and Jones, M.N. (1989) The characterization of liposomes with covalently attached proteins. Biochim. Biophys. Acta 978,17-24. 

Chappell, J. D., Prota, A. E., Dermody, T. S., and Stehle, T. (2002). Crystal structure of reovirus attachment protein sigmal reveals evolutionary relationship to adenovirus fiber. EMBOJ. 21, 1-11. 

Of the extremely diverse examples of protein modifications observed in eukaryotic cells, the modifications by lipid (and glycolipid) molecules are of special interest because lipid-attached proteins can be anchored at the membrane, although all of these proteins are not always anchored. So far, three groups of membrane anchoring proteins have been noted (Fig. 5). 

Huang WJ, Taylor S, Fu KF, Lin Y, Zhang DH, Hanks TW, Rao AM, Sun YP (2002) Attaching proteins to carbon nanotubes via diimide-activated amidation. Nano Lett. 2 311-314. 

As detailed in chapter 17, biological membranes are basically lipid—think fat or oil—in nature with some attached proteins. As such, these thin sheets of phospholipids and proteins are nearly impermeable to charged particles such as sodium, potassium, or chloride ions. While the isolation of the cell interior from the exterior ionic environment is critical in many ways, it is also true that controlled permeability to ions may be critical. In fact, it is the near-impermeability of biological membranes to ions that permits control of ion transport across them by certain, specific proteins. 

Special tasks. Some lipids have adopted special roles in the body. Steroids, eicosanoids, and some metabolites of phospholipids have signaling functions. They serve as hormones, mediators, and second messengers (see p.370). Other lipids form anchors to attach proteins to membranes (see p.214). The lipids also produce cofactors for enzymatic reactions—e.g., vitamin K (see p.52) and ubiquinone (see p.l04). The carotenoid retinal, a light-sensitive lipid, is of central importance in the process of vision (see p.358). 

Therefore the preparation of an activated, preformed polymer which was capable of reacting directly with the protein molecules in solution provided a less traumatic one-step incorporation of the protein into the polymer. This new method retained the biological activity of the attached protein and was more reproducible. As one possible application of our technology, the development of a novel immunoassay is also described here. 

Our proposal is not theoretical. Researchers have used reticulated hydrophobic polyurethanes as liver assist devices with some success. We will discuss this research and future work in detail later. For now, it is useful to present an overview. Matsushita et al. inoculated a reticulated polyurethane with porcine hepatic cells, "fhe device functioned as noted, but it was necessary to separate the plasma from the blood because conventional hydrophobic polyurethanes are not hemocompatible. In addition, the technique made no provision for cell attachment. Workers in our laboratory grafted a hydrophilic polyurethane to the structural members of a hydrophobic reticulated foam in an effort to make the composite hemocompatible. Additionally, this gave us the opportunity to add cell attachment proteins. 

Prenylation (covalent attachment of an isoprenoid see Fig. 27-30) is a common mechanism by which proteins are anchored to the inner surface of cellular membranes in mammals (see Fig. 11-14). In some of these proteins the attached lipid is the 15-carbon farnesyl group others have the 20-carbon geranylgeranyl group. Different enzymes attach the two types of lipids. It is possible that prenylation reactions target proteins to different membranes, depending on which lipid is attached. Protein prenylation is another important role for the isoprene derivatives of the pathway to cholesterol. 

FIGURE 27-41 Three-step cascade pathway by which ubiquitin is attached to a protein. Two different enzyme-ubiquitin intermediates are involved. The free carboxyl group of ubiquitin s carboxyl-terminal Gly residue is ultimately linked through an amide (isopeptide) bond to an e-amino group of a Lys residue of the target protein. Additional cycles produce polyubiquitin, a covalent polymer of ubiquitin subunits that targets the attached protein for destruction in eukaryotes. 

Presynaptic nerve terminals may contain as few as a hundred vesicles, which must be recycled rapidly after exocytosis in order to allow for repetitive fir-jHg 558,559,583 gevera proteins are needed for endocyto-sis (step 7 in Fig. 30-20). These include endophilin I,584 the vesicle transport ATPase NSF,574 GTPases,565 and the soluble NSF attachment protein a-SNAP (which is not related to SNAP-25).585... 

An attractive strategy to improve CNS drug delivery is to link a nontransportable drug with a vector to the BBB. These moieties can work as molecular Trojan horses to transport across the BBB attached proteins, DNA molecules, and drug micro- and nanocarriers facilitating their penetration through the BBB. The choice of a vector moiety and a type of a linker is crucial for the success of this method of drug delivery. 

Probably the best means of protein attachment is through highly specific affinity interactions (e.g., streptavidin-biotin or His-tag-nickel-chelates) [111]. Proteins fused with a high-affinity tag, at their amino or carboxyl terminus, are finked to the surface of the chip via this tag, and hence all the attached proteins should orient uniformly away from the surface. Using this method, immobilized proteins are more likely to remain in their native conformation, while the analytes have easier access to the active sites of the proteins. 

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