Proteins transferred

High-density lipoproteins (HDL) have much longer life spans in the body (5 to 6 days) than other lipoproteins. Newly formed HDL contains virtually no cholesterol ester. However, over time, cholesterol esters are accumulated through the action of lecithin cholesterol acyltransferase (LCAT), a 59-kD glycoprotein associated with HDLs. Another associated protein, cholesterol ester transfer protein, transfers some of these esters to VLDL and LDL. Alternatively, HDLs function to return cholesterol and cholesterol esters to the liver. This latter process apparently explains the correlation between high HDL levels and reduced risk of cardiovascular disease. (High LDL levels, on the other hand, are correlated with an increased risk of coronary artery and cardiovascular disease.)... 

Under physiological conditions, transfer of Et3PAufrom hemoglobin to albumin has been observed. The transfer is direct and does not require a low-molecular weight thiol such as GSH as an intermediary and indicates that inter-protein transfer of gold can occur spontaneously. The rapid and efficient manner of establishing this equilibrium (for transauration ) could determine many ofthe effects of intracellular and extracellular chemistry of gold. 

With acid or basic substrates protein transfer may contribute to recognition by the model receptor, but binding to lactams involves hydrogen bonds exclusively. Recent research321 has concluded that hydrogen bonds are not so often created as they are merely exchanged in aqueous solutions, and the overall enthalpic changes are modest. 

Szewczyk B, Summers DF (1987) Fluorescent staining of proteins transferred to nitrocellulose allowing for subsequent probing with antisera. Anal Biochem 164 303-306... 

NAD PI I gives up hydrogen atoms to the flavo protein NADPH— cytochrome P450 reductase and becomes NADP+. The reduced flavo protein transfers these reducing equivalents to cytochrome P450. The reducing... 

Hydration of phospholipid head groups is essential properties not only for stabilizing bilayer structures in an aqueous environment, but also for fusion or endocytosis of biological membranes including protein transfers [33-35]. Hydration or swelling behavior has only been studied by indirect methods such as X-ray diffraction [36], differential scanning calorimetry (DSC) [37], and H-NMR [38,39]. 

Knudsen KA. 1985. Proteins transferred to nitrocellulose for use as immunogens. Anal Biochem 147 285-288. 

The N2-fixing enzyme used by the bacteria is nitrogenase. It consists of two components an Fe protein that contains an [Fe4S4] cluster as a redox system (see p. 106), accepts electrons from ferredoxin, and donates them to the second component, the Fe-Mo protein. This molybdenum-containing protein transfers the electrons to N2 and thus, via various intermediate steps, produces ammonia (NH3). Some of the reducing equivalents are transferred in a side-reaction to In addition to NH3, hydrogen is therefore always produced as well. 

G proteins transfer signals from 7-helix receptors to effector proteins (see above). G protein are heterotrimers consisting of three different types of subunit (a, 3, and y see p.224). The a-subunit can bind GDP or GTP (hence the name G protein ) and has GTPase activity. Receptor-coupled G proteins are related to other GTP-binding proteins such as Ras (see pp. 388, 398) and EF-Tu (see p. 252). 

Protein transfer from the brain into CSF, and from blood into CSF, follows the law of diffusion as a function of molecular size. The diffusion-related transfer of proteins into CSF is the cause for molecular size-dependent discrimination (i.e., selectivity) of the barrier function. As a consequence, we have larger CSF/ serum quotients for the smaller molecules QAlb > QIgG > QIgA > QIgM. Again, the smaller albumin molecule equilibrates faster between blood and CSF than do the larger molecules of IgG, IgA, or IgM. 

The membrane is rinsed with H2O after protein transfer (electrotransfer or dot blot) and dipped to methanol for some seconds. Immediately after this (avoid drying), the membrane is agitated in Soln. A for maximum 2 min and destained by several changes of Soln. B. When the membrane is dry, blue bands are visible on a slight blue background. Membranes stained by this method are not suitable for any immunochemical detection. 

Molecular (+)-brefeldin polyket./inhibitor of protein transfer from... 

Rate of protein transfer to or from a reverse micellar phase and factors affecting the rate are important for the practical applications of RME for the extraction and purification of proteins/enzymes and for scale-up. The mechanism of protein exchange between two immiscible phases (Fig. 2) can be divided into three steps [36] the diffusion of protein from bulk aqueous solution to the interface, the formation of a protein-containing micelle at the interface, and the diffusion of a protein-containing micelle in to the organic phase. The reverse steps are applicable for back transfer with the coalescence of protein-filled RM with the interface to release the protein. The overall mass transfer rate during an extraction processes will depend on which of these steps is rate limiting. 

Efficient extraction of proteins has been reported with reverse micellar liquid membrane systems, where the pores of the membrane are filled with the reverse micellar phase and the enzyme is extracted from the aqueous phase on one side of membrane while the back extraction into a second aqueous phase takes place at the other side. By this, both the forward and back extractions can be performed using one membrane module [132,208]. Armstrong and Li [209] confirmed the general trends observed in phase transfer using a glass diffusion cell with a reverse micellar liquid membrane. Electrostatic interactions and surfactant concentration affected the protein transfer into the organic membrane and... 

VirB proteins in the secretion system most form the membrane channel or serve as ATPases to provide energy for channel assembly or for export processes. Several of these VirB proteins form the T-pilus. The function of the T-pilus remains unclear it may be required as the channel for T-DNA and Vir protein transfer, or it may act as a hook to grab hold of the recipient host cell and bring plant and bacterium close together for efficient gene transfer (Gelvin, 2003, LaCroix et ah, 2006a). 

When you are considering catalysis by amino acids, you speak essentially of what is called the physical adsorption, where the energy of interaction between the catalyst and the adsorbed species is of the order of Van der Waals forces that can be sufficient for protein transfer and for other charge transfers that seem to play a very important role in biological reactions but are hardly sufficient for other reactions. 

Synthesis of a Selective, Non-Peptide, Non-Sulfhydryl Farnesyl Protein Transfer Inhibitor (Antitumor Agent)... 

Schering Plough demonstrated the kinetic resolution of a secondary amine (24) via enzyme-catalyzed acylation of a pendant piperidine (Scheme 7.13) [32]. The compound 27 is a selective, non-peptide, non-sulfhydryl farnesyl protein transfer inhibitor undergoing clinical trials as a antitumor agent for the treatment of solid tumors. The racemic substrate (24) does not contain a chiral center but exists as a pair of enantiomers due to atropisomerism about the exocylic double bond. The lipase Toyobo LIP-300 (lipoprotein lipase from Ps. aeruginosa) catalyzed the isobu-tylation of the (+) enantiomer (26), with MTBE as solvent and 2,2,2-trifluoroethyl isobutyrate as acyl donor [32]. The acylation of racemic 24 yielded (+) 26 at 97% and (-) 25 at 96.3% after 24h with an E >200. The undesired enantiomer (25)... 

Scheme 7.13 Enzymatic resolution for the synthesis of 27 for the sythesis of selective, non-peptide, non-sulfhydryl farnesyl protein transfer inhibitor currently undergoing phase II clinical trials for the treatment of solid tumors.

Cholesteryl ester Cholesteryl ester transfer protein transfer protein... 

Salinovich, O and Montelaro, R. C. (1986) Reversible staining and peptide mapping of proteins transferred to nitrocellulose after separations by sodium dodceylsulfate-polyacrylamide gel electrophoresis Anal Biochem 156,341-347. 

In some cases, stained blots are used only to identify protein band patterns while leaving the gel unmodified for subsequent steps (UNITB3.3). If such minimal protein transfer is desired, contact blotting is a suitable alternative. This unit also describes procedures for eluting proteins from membranes using detergents (Basic Protocol 2) or acidic extraction with organic solvents (Alternate Protocol 4). 

The protocols listed in this unit should be sufficient for the transfer of most proteins. The efficiency of protein transfer is easily assessed by staining both the transfer membrane and the gel after transfer. When difficulty is encountered in the transfer recovery of a specific protein, several factors can be considered to improve transfer efficiency. 

Another problem often encountered with poor transfer of proteins relates to the amount of SDS in the gel, which coats the proteins and improves mobility during electrotransfer. Nitrocellulose membranes are not sensitive to the amount of SDS in the gel and can even tolerate addition of SDS to the transfer buffer to encourage transfer of recalcitrant proteins. PVDF membranes are more sensitive to excess SDS, which can inhibit protein binding to the hydro-phobic membrane nevertheless, in a limited number of cases, addition of SDS to the transfer buffer may be beneficial. Low-retention PVDF membranes are particularly sensitive to excess SDS concentrations, whereas high-retention PVDF membranes are less so. Methanol is a common component of many transfer buffers because it facilitates dissociation of bound SDS from proteins. Therefore, if proteins transfer from the gel efficiently but do not bind well to PVDF membranes, the methanol concentration can be increased to 20% and/or the gel can be preequilibrated in transfer buffer for 15 to 30 min prior to transfer to reduce the SDS concen-... 

---