Interaction membrane-substrate

Magnetic resonance techniques have again been popular for studying enzymes which are involved in phosphate hydrolysis and transfer. 31P or 19F N.m.r.1-2 and spinlabelling3 have all been used to study the interaction of substrates with these enzymes, while affinity labelling4 5 6 7 is another technique which has been used to obtain information about the sequence and conformation of amino-acid chains at the active sites of enzymes. Recently, these experimental methods have been applied to the study of cell membranes,6-7 and these are mentioned in a new series of books concerned with enzymes in biological membranes.8 A new journal, Trends in Biochemical Sciences, which contains concise, up-to-date reviews on these and other topics is published by Elsevier on behalf of the International Union of Biochemistry. 

Murakami et al. studied alternative pyridoxamine-surfactant systems [23]. These authors synthesized hydrophobic pyridoxamine derivatives (30 and 31) and peptide lipid molecules (32-35). Catalyst 30 or 31 and the peptide lipids formed bilayer membranes in water, which showed transamination reactivity in the presence of metal ions such as Cu(ii). It was proposed that the pyridoxamine moiety was placed in the so-called hydrogen-belt domain interposed between the polar surface region and the hydrophobic domain that is composed of double-chain segments within the bilayer assembly. The basic group (such as imidazole) in the peptide lipid molecules could catalyze the proton transfer involved in the transamination reaction. In addition, marked substrate discrimination by these bilayer membrane systems was performed through hydrophobic interactions between substrates and the catalytic site. 

The question of how DnaJ proteins interact with substrates and mediate their transfer onto Hsp70 partner proteins is not answered for any of the three classes of DnaJ proteins. Some DnaJ homologs have broad substrate specificity, such as E. coli DnaJ and yeast Ydjl, while others have more restricted substrate spectra. In particular the DnaJ proteins of class III may either bind a restricted number of substrates, such as the clathrin-specific auxilin or the kinesin light chain, or they may not bind substrates themselves but rather are positioned in close proximity to substrates. The latter seems to be the case for Dj 1A in the plasma membrane of E. coli (Clarke et al, 1997 Kelley and Georgopoulos, 1997a), Sec63 at the translocation pore in the ER (Corsi and Schekman, 1996 Rapoport et al., 1996), and cysteine string proteins on the surface of neurosecretory vesicles (Buchner and Bundersen, 1997). 

An intuitive feeling for the phenomenon of countertransport can be appreciated as follows the two substrates S and P compete with each other for combination with the free carrier E. At the face at which P is at the high concentration, it will compete more effectively with S than it will at the other face. Hence less S will flow from the face at which most P flows. There will be thus a net flow of S in a direction opposite to the flow of P. All this occurs because, on the carrier model, the interaction of substrates and carrier at one face of the membrane are separated from the interactions at the other face by the measurable activation energy barrier for the interconversion of the two forms of the free carrier. 

Much of the literature uses simpler synthetic mimics of cell membranes, consisting of vesicles (also known as liposomes) and occasionally flat bilayers supported on a substrate. Most contain no protein in order to focus on polymer-lipid interactions. Membranes meant to mimic mammalian membranes generally consist of PC and may include some cholesterol. Red blood cells (RBCs) also serve as a model system, as they do not divide. Gram-positive bacteria mimics generally consist of PC and cardiolipin, while gram-negative bacteria mimics have PE and PC. 

It must be pointed out that yeast-mediated biotransformations may be complicated by side reactions that interfere or even dominate the desired conversion workup may be somewhat time consuming and messy due to the separation of the product from the huge amount of biomass. The ideal interactions between the yeast and the substrate are scarcely found in praxi. Ideal interactions between substrate yeast and product include that both substrate and product are able to pass the cell membrane, should be soluble in the fermentation medium, and must not inactivate the catalytic activity of the involved microbial enzymes. Finally, high turnover rates and high regio- and stereospecificity are also of high practical relevance. Some advice on how to deal with these basic problems often encountered in such biotransformations is provided in Fig. 1 [9,11,13,19,20]. 

The interest in vesicles as models for cell biomembranes has led to much work on the interactions within and between lipid layers. The primary contributions to vesicle stability and curvature include those familiar to us already, the electrostatic interactions between charged head groups (Chapter V) and the van der Waals interaction between layers (Chapter VI). An additional force due to thermal fluctuations in membranes produces a steric repulsion between membranes known as the Helfrich or undulation interaction. This force has been quantified by Sackmann and co-workers using reflection interference contrast microscopy to monitor vesicles weakly adhering to a solid substrate [78]. Membrane fluctuation forces may influence the interactions between proteins embedded in them [79]. Finally, in balance with these forces, bending elasticity helps determine shape transitions [80], interactions between inclusions [81], aggregation of membrane junctions [82], and unbinding of pinched membranes [83]. Specific interactions between membrane embedded receptors add an additional complication to biomembrane behavior. These have been stud-... 

In biological systems molecular assemblies connected by non-covalent interactions are as common as biopolymers. Examples arc protein and DNA helices, enzyme-substrate and multienzyme complexes, bilayer lipid membranes (BLMs), and aggregates of biopolymers forming various aqueous gels, e.g, the eye lens. About 50% of the organic substances in humans are accounted for by the membrane structures of cells, which constitute the medium for the vast majority of biochemical reactions. Evidently organic synthesis should also develop tools to mimic the Structure and propertiesof biopolymer, biomembrane, and gel structures in aqueous media. 

Tyrosine phosphorylated IRS interacts with and activates PI 3-kinase [3]. Binding takes place via the SRC homology 2 (SH2) domain of the PI 3-kinase regulatory subunit. The resulting complex consisting of INSR, IRS, and PI 3-kinase facilitates interaction of the activated PI 3-kinase catalytic subunit with the phospholipid substrates in the plasma membrane. Generation of PI 3-phosphates in the plasma membrane reemits phospholipid dependent kinases (PDKl and PDK2) which subsequently phosphorylate and activate the serine/threonine kinase Akt (synonym protein... 

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