Lipase specificity

The mobilization of stored fat requires the hydrolytic release of fatty adds and glycerol from their TAG form. This process is initiated by hormone-sensitive lipase, which removes a fatty acid from carbon 1 and/or carbon 3 of the TAG. Additional lipases specific for diacylglyc-erol or monoacylglycerol remove the remaining fatty acids. 

Specificity. A study of lipase specificity requires that the enzyme and substrates be virtually pure. Contamination of the lipase preparation with esterases gives rise to misleading results. Pure, synthetic substrates of known configuration are essential, and the same available surface area should be present after emulsification for meaningful data to be obtained. Since most of the earlier workers disregarded one or more of these variables, their data will not be included here. 

Gas-Liquid Chromatography. Gas-liquid chromatography (GLC) affords both a qualitative and, if adequate internal standards are used, a quantitative analysis of the products of lipolysis. It is necessary, however, first to isolate the acids by a suitable method and then to inject them as free acids or as esters. The partial glycerides can be isolated by thin-layer chromatography and can also be determined by GLC of suitable derivatives. The acid(s) remaining in the partial glycerides can be identified readily by GLC following transesterification. Jensen and co-workers have utilized these techniques in their studies of lipase specificity (Jensen et al 1964). 

Cooke, B.C. 1973. The influence of incubation temperature on microbial lipase specificity. N.Z. J. Dairy Sci. Technol. 8, 126. 

Villeneuve, P., Pina, M., and Graille, J. 1996. Determination of pregastric lipase specificity in young ruminants. Chem. Phys. Lipids, 83,161-168. 

Figure 2-7 Plane of Symmetry of a Glycerol Molecule (Top) and Mirror Image of Two Enantiomers of a Mono-Acylglycerol (bottom). Source Reprinted with permission from P. Ville-neuve and T.A. Foglia, Lipase Specificities Potential Application in Bioconversions, Inform, 8, pp. 640-650, 1997, AOCS Press.

Table 10-6 Examples of Lipase Specificities Specificity Lipase...

Lipase catalysis constitutes an alternative to the method of interesterification using chemical catalysts. Products of different physical properties may be obtained, according to lipase specificity (99,100). 

Microbial lipases may be used as biocatalysts for interesterification. Lipase-assisted interesterification offers possibilities for transformation of lipids beyond those possible using chemical interesterification (11). Enzymatic interesterihcation has several advantages over the chemical-assisted reactions, such as mild reaction conditions leading to reduced energy consumption and less thermal damage to reactants and products, the possibility of lipase specificity toward their natural substrates, as well as high catalytic efficiency. Lipase-catalyzed interesterihcation reactions, in contrast to those carried out with chemical catalysts, yield different types of products depending on the specihcity of the lipase used. 

Specificity is one of the most striking properties of enzyme molecules. Enzyme specificity can be defined as a comparative difference in rates of catalysis of certain reactions. After an enzyme is identified as a hpase, several specificities within the class are identified or can be expected to occur. The main advantage of lipases, which differentiate enzymatic reactions from chemically-catalyzed reactions, is lipase specificity. Lipases have turned out to be very useful enzymes for catalyzing various types of reactions with a rather wide substrate specificity. The fatty acid specificity of lipases has been exploited to produce structured lipids and to enrich lipids with specific fatty acids to improve the nutritional characteristics of lipids (24). Certain lipases display positional specificity (regiospecificity) toward fatty acyl groups in a TAG molecule as well as fatty acid selectivity. 

Short-chain fatty acid preference also is a lipase specificity that can be exploited on an industrial scale. Such lipases may be used in the production of low-calorie structured lipids or in the dairy industry to obtain specific flavor components by the release of short- or medium-chain fatty acids from milkfat (25). 

Villeneuve, P. Foglia, T.A. Lipase specificities potential application in lipid bioconversions. Inform 1997, 8 (6), 640-651. 

Ke, T., Tidor, B., and Klibanov, A. M., Molecular-modeling calculations for enzymatic enantioselectivity taking hydration into account, Biotechnol. Bioeng., 57,741-745,1998. Haeffner, F., Norin, T., and Hull, K., Molecular modeling of the enantioselectivity in lipase-catalyzed transesterification reactions, Biophys. J., 74, 1251-1262, 1998. Bernstein, F. C., Koetzle, T. R, WiUiams, G. J. B., Meyer, E. F. J., Brice, M. D., Rodgers, J. R., Kennard, O., Shimanouchi, T., and Tasumi, M., The protein data bank a computer-based archival file for macromolecular structures, J. Mol. Biol., 112, 535-542, 1977. Parida, S. and Dordick, J. S., Tailoring lipase specificity by solvent and substrate chemistries, J. Org. Chem., 58, 3238-3244, 1993. 

Ergan, F. and Trani, M., Effect of lipase specificity on triglyceride synthesis, Biotechnol. Lett., 13, 19-24, 1991. 

Lipases specific for a particular class of fatty acids Geotrichum candidum has been found specific for omega-9 fatty acids others are specific for short-chain fatty acids, and still others for long-chain fatty acids... 

One interesting technology uses lipases in the form of cross-linked enzyme crystals (CLECs) (Margolin 1996). This immobilization method does not use any solid support and the lipase specific activity (units of activity/g of immobilized catalyst) of the immobilized lipase derivative can be enhanced by 10-fold because there is no inert support, that usually represent more than 90% of the catalyst weight in the case of carrier-bound enzymes. These cross-linked crystals have been used for the chiral resolution of commercially important organic compounds, such as ibuprofen. 

Lipases have several attractive properties that make them suitable for several applications. Lipase specificity eliminates the production of undesired products in the waste stream, decreases side reactions, and simplifies separation problems (Hasan et al., 2006 Pandey et al., 1999). Moreover, lipase reactions can be carried out under mild pH and temperature conditions, which reduces energy consumption and protects unstable substrates and products from denaturation. Another potential feature of lipases is their stability in an organic solvent, where they catalyze reactions... 

As reported elsewhere in this volume, a lipase, specifically located in the bran tissues of wheat grain, is responsible for the hydrolysis of endogenous triacylglycerols (TG) in wholemeal to free (unesterified) fatty acids, 60-65% of which are polyunsaturated fatty acids (F-puFA). This process occurs in wholemeal flour during storage under ambient conditions. 

Jensen, RG, FA Dejong, RM Clark. (1983). Determination of lipase specificity. Lipids 18 239-252. 

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