Immobilization phospholipase

Choline-containing phospholipids are determined by co-immobilizing phospholipase D and choline oxidase onto a platinum electrode ... 

L.R. Juneja, N. Hibi, T. Yamane, and S. Shimizu Repeated batch and continuous operations for phosphatidylglycerol synthesis from phosphatidylcholine with immobilized phospholipase U. Applied Microbiology and Biotechnology 27 (1987) 146-151. 

N. Dittrich and R.U. Hofmann Transphosphatidylation by immobilized phospholipase D in aqneons media. Biotechnology and Applied Biochemistry 34 (2001) 189-194. 

Fig. 1.2. (a) Scheme for enzymatic determination of phosphatidyl choline (PC) using phospholipase C, alkaline phosphatase and choline oxidase, (b) Schematic diagram of the flow injection manifold including a co-immobilized phospholipase C-alkaline phosphatase-choline oxidase immobilized enzyme reactor (PLC-AP-ChO IMER) for the conversion of phosphatidyl choline to hydrogen peroxide. The hydrogen peroxide produced was detected using a chemiluminescence detector. 

Anthonsen T, D Arrigo P, Pedrocchi-Fantoni G, Secundo F, Servi S, Sundby E. 1999. Phospholipids hydrolysis in organic solvents catalyzed by immobilized phospholipase C. J Mol Catal B Enzym 6 125-132. 

There is evidence for immunosuppressive effects of PAHs in rodents (Davila et al. 1997). For example, strong immunosuppressive effects were reported in mice that had been dosed with benzo[fl]pyrene and 3-methyl cholanthrene, effects that persisted for up to 18 months (Environmental Health Criteria 202). Multiple immu-notoxic effects have been reported in rodents, and there is evidence that these result from disturbance of calcium homeostasis (Davila et al. 1997). PAHs can activate protein tyrosine kinases in T cells that initiate the activation of a form of phospholipase C. Consequently, release of inositol triphosphate—a molecule that immobilizes Ca + from storage pools in the endoplasmic reticulum—is enhanced. 

Fig. 4.12(a). An outline structure of a protein (here the enzyme phospholipase A2), showing a-helical runs of amino acids as cylinders (A-E) and anti-parallel P-sheet runs as heavy black arrows. Disulfide cross-links are shown (the enzyme is extracellular), and runs of no a/p secondary structure appear as thin lines. The structure is relatively immobile, and binds calcium in a constrained loop. (Reproduced with permission from Professor J. Drenth.)... 

Choline-containing phospholipids have been determined in human serum using an IMER consisting of coimmobilized phospholipase D and choline oxidase [51]. Recently, immobilized glutamate oxidase was used to determine L-glutamic acid in culture media [52],... 

Phospholipases must have a similar mechanism of orientation at the interface because their substrates are also immobilized (except for lateral movements) in a supersubstrate structure. The enzyme must attach itself to the supersubstrate in the correct orientation in order to place the reactive center toward the substrate (Figure 7). In pancreatic phospholipase and many other phospholipases the binding to the supersubstrate may be electrostatic rather than hydrophobic so that the enzyme has an electrostatic head. Supersubstrate binding sites of mixed type or hydrogen bonding sites may also exist. 

Active immobilized enzyme isolation of specific anti-phospholipase antibodies Study of the kinetics and stability of the immobilized enzyme and its potential use as a probe for membrane surface phospholipids... 

A porous vessel filled with the biocatalyst and immersed in recirculating toluene was applied for the preparation of methyl trans-(2 J ,3S)-(4-methoxyphenyl)glyddate (an intermediate of Diltiazem) from its racemic methyl ester [88]. In this system, hydrolysis of the ester with Lecitase Ultra (a commercially available phospholipase) immobilized in macroporous gelatin organo-gel (gelozyme) gave the desired product in 47% yield and >99% ee. 

CIAP is usually prepared from unweaned calf intestinal mucosa. The membrane-bound APase can be selectively released (up to 90%) by treatment with bacterial (S. aureus) phosphatidylinositol-specific phospholipase C (20). Affinity chromatography using a Sepharose-immobilized triazine dye analog that carries a phosphonate-terminal aminobenzene ring yields a 330-fold one-step purification of CIAP from a crude intestinal extract (21). The purity of CIAP, which can be specifically eluted from the column with a 5 mM Pj solution, is reported to be equivalent to that of a commercial high purity preparation. 

Finally, any enzymatic process using lipases or phospholipases can be triggered at will by changing the internal balance between the core water and the free water. On the shelf, the product will contain enzymes that will be immobilized and temporarily deactivated by the bound water, and when the microemulsion is further diluted, during use, the enzymes will become free and active and the reaction will be triggered. 

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