Lipases ester bond hydrolysis

Lipases are the enzymes for which a number of examples of a promiscuous activity have been reported. Thus, in addition to their original activity comprising hydrolysis of lipids and, generally, catalysis of the hydrolysis or formation of carboxylic esters [107], lipases have been found to catalyze not only the carbon-nitrogen bond hydrolysis/formation (in this case, acting as proteases) but also the carbon-carbon bond-forming reactions. The first example of a lipase-catalyzed Michael addition to 2-(trifluoromethyl)propenoic acid was described as early as in 1986 [108]. Michael addition of secondary amines to acrylonitrile is up to 100-fold faster in the presence of various preparations of the hpase from Candida antariica (CAL-B) than in the absence of a biocatalyst (Scheme 5.20) [109]. 

Effect of Molecular Weight of Polyester on the Hydrolysis by Rhizopus lipase. Using three kinds of polyesters, PCL-diol (I), polyhexameth-ylene adipate (II), and a copolyester (ill) made from 1,6-hexamethyl-enediol and a 70 30 molar ratio mixture of e- caprolactone and adipic acid, the effects of the of polyester on the hydrolysis by lipase were examined (Figure k) Mn did not affect the rates of hydrolysis by R. arrhizus and delemar lipases when Vln was more than about UOOO. This would indicate these lipases randomly splits ester bonds in pol-mer chains. In contrast, when TEi was less than about i4000 2 the rates of the enzymatic hydrolysis were faster with the smaller Mn of polyesters. This corresponded to the fact that Tm was lower with the smaller Mn of polyesters. 

Hydrolysis of Copolyamide-esters (CPAEs) by Lipase (jj,). CPAEs were synthesized by the amide-ester interchange reaction between polyamide and polyester. The length of the polyamide blocks was measured after hydrolysis of ester bonds in CPAE by alkali at 30 C. The infrared spectra after hydrolyzing ester bonds on CPAEs showed that the ester bonds were almost completely removed. The molecular weight distribution of polyamide blocks was examined by GPC (Table II). The following samples were used CPAE-1 (reaction time for synthesis, 1 hr) and CPAE-2 (reaction time, U hr) composed of nylon 6 and PCL at a 50/50 molar ratio, CPAE-3 (reaction time, 1 hr) and CPAE-U (reaction time,... 

The stability of the ester surfactants against enzymatic hydrolysis by two different microbial Upases, Mucor miehei lipase (MML) and Candida antarc-tica lipase B (CALB) added separately to the surfactant solutions, was also investigated, see Fig. 5 [19]. It is obvious that hydrolysis of the unsubstituted surfactant is much faster with both CALB and MML than that of the substituted surfactants, i.e., increased steric hindrance near the ester bond leads to decreased hydrolysis rate. Since the specificity of the enzyme against its substrate is determined by the structure of the active site, it can be concluded, as expected, that the straight chain surfactant most easily fits into the active site of both enzymes. 

Hydrolases, which catalyze the hydrolysis of various bonds. The best-known subcategory of hydrolases are the lipases, which hydrolyze ester bonds. In the example of human pancreatic lipase, which is the main enzyme responsible for breaking down fats in the human digestive system, a lipase acts to convert triglyceride substrates found in oils from food to monoglycerides and free fatty acids. In the chemical industry, lipases are also used, for instance, to catalyze the —C N —CONH2 reaction, for the synthesis of acrylamide from acrylonitril, or nicotinic acid from 3-pyridylnitrile. 

The overall process for this enzymatic resolution is compared with the conventional chemical process in Fig. 14. The enzymatic process can skip several tedious steps which are necessary in chemical resolution and this is a considerable practical advantage. There have been several reports on the application of enzymatic asymmetric hydrolysis to the optical resolution of pantolactone [141, 142], In these cases, esterified substrates, such as O-acetyl or O-formyl pantolactone, and lipases were used as the starting materials and catalysts, respectively. Since the lactonase of F. oxysporum hydrolyzes the intramolecular ester bond of pantolactone, it is not necessary to modify the substrate, pantolactone. This is one of the practical advantages of this enzyme. 

The use of extracellular lipases of microbial origin to catalyze the stereoselective hydrolysis of esters of 3-acylthio-2-methylpropionic acid in an aqueous system has been demonstrated to produce optically active 3-acylthio-2-methyl-propionic acid [41-43], The synthesis of the chiral side chain of captopril by the lipase-catalyzed enantioselective hydrolysis of the thioester bond of racemic 3-acetylthio-2-methylpropionic acid (15) to yield 5 -(-)-(15) has been demonstrated [44], Among various lipases evaluated, lipase from Rhizopus oryzae ATCC 24563 (heat-dried cells), BMS lipase (extracellular lipase derived from the fermentation of Pseudomonas sp. SC 13856), and lipase PS-30 from Pseudomonas cepacia in an organic solvent system (l,l,2-trichloro-l,2,2-tri-fluoroethane or toluene) catalyzed the hydrolysis of thioester bond of undesired enantiomer of racemic (15) to yield desired S-(-) (15), R-(+)-3-mercapto-2-methylpropionic acid (16) and acetic acid (17) (Fig. 8A). The reaction yield of... 

Lipoprotein lipase (EC 3.1.1.34) is an enzyme or group of enzymes which catalyze the hydrolysis of the 1(3) ester bond(s) of triacylglycerols and the 1 ester bond of phospholipids. The enzyme plays a central role in lipoprotein metabolism, being responsible in particular for the hydrolysis of chylomicron and VLDL triglycerides and the formation of remnant particles from these lipoproteins. There have been reviews of this enzyme [e.g., (N9, Ql)] and lipoprotein lipase will not be discussed in detail in this review. Familial lipoprotein lipase deficiency and related disorders of chylomicron metabolism have also been reviewed (B58, N8) and will not be discussed in detail. 

Lipases are a special class of esterases that also catalyze the hydrolytic cleavage of ester bonds, but differ in their substrate spectrum. Lipases have the special capability to catalyze the hydrolysis of water-insoluble substrates such as fats and lipids. Like many other enzyme-catalyzed reactions, the ester hydrolysis is a reversible process, which allows using lipases and other esterases for the synthesis of esters. The use of lipases as catalysts in synthetic chemistry is described elsewhere in this chapter. 

Animals store energy in the form of triacylglycerols, kept in a layer of fat cells below the surface of the skin. This fat serves to insulate the organism, as well as provide energy for its metabolic needs for long periods. The first step in the metabolism of a triacylglycerol is hydrolysis of the ester bonds to form glycerol and three fatty acids. This reaction is simply ester hydrolysis. In cells, this reaction is carried out with enzymes called lipases. 

Lipases are widely distributed among animals, plants, and microorganisms. They catalyze the hydrolysis of ester bonds at the fat/water interface. The mechanism of the lipase-catalyzed hydrolysis of esters involves two consecutive steps [51]. First, the ester is attacked by a nucleophilic group of the enzyme - normally an OH of a serine or a SH of a cysteine - to form an acyl-enzyme intermediate with concomitant liberation of the alcohol moiety of the ester. The acyl-enzyme is subsequently attacked by a water molecule to give the product. 

Lipases, developed by nature for the hydrolysis of fatty acyl ester bonds, have also been explored for the hydrolysis of fats and oils. However, because of issues of cost, stability, and productivity, they are not presently employed in industrial lipid hydrolysis. 

In general terms, the crystallographic results show that lipases contain several distinct sites, each responsible for a specific function. The hydrolysis of the ester bond is accomplished by the catalytic triad, responsible for nucleophilic attack on the carbonyl carbon of the scissile ester bond, assisted by the oxyanion hole, which stabilizes the tetrahedral intermediates. The fatty acid recognition pocket defines the specificity of the leaving acid. There is also one or more interface activation sites, responsible for the conformational change in the enzyme. In this section the discussion is on the available structural data relevant to the function of all these sites. 

Acetylcholinesterase (AChE) catalyses the hydrolysis of the ester bond of acetylcholine to yield choline and acetate (Sussman et al., 1991). This is a critical reaction for the termination of impulses transmitted through cholinergic synapses. It is a highly efficient catalyst, with reaction rates approaching the diffusion limit. Its overall structure resembles the lipases with an active site gorge. Above the base of the gorge is the reactive serine to be activated by the classical (Ser-200...His-440...Glu-327) catalytic triad. 

Developments in this class of enzymes are so rapidly and promising that they are likely to be a strong influence in the food industry. Examples are the production of entirely new foods and the production of a wide variety of effective flavours or emulsifiers. Under aqueous conditions lipases catalyse the breakdown (hydrolysis) of fats and other ester bond containing compounds. Moreover, lipases appear to be effective... 

The considerations in the previous section need to be addressed and customized for every screening project. As an example, when we set out to develop a new library of esterases for synthetic chemistry use, we first needed to determine the criteria that would be used in the screening project. Esterases and lipases catalyze the hydrolysis of ester bonds as shown in Scheme 1 and are useful for reactions requiring different regioselectivities, chemoselectivities, and stereoselectivities depending on the enzyme s substrate specificities. 

Several types of enzymes have found uses in LADD compositions [4,48], Most common are proteases, amylases, and lipases, which attack proteinaceous, starchy, and fatty soils, respectively. Proteases work by hydrolyzing peptide bonds in proteins. Proteases differ in their specificity toward peptide bonds. The typical protease used in LADD formulations, bacterial alkaline protease (subtilisin), is very nonspecific. That is, it will attack all types of peptide bonds in proteins. In contrast to proteases, amylases catalyze the hydrolysis of starch. They attack the internal ether bonds between glucose units, yielding shorter, water-soluble chains called dextrins. Lipases work by hydrolyzing the ester bonds in fats and oils. Often, combinations are used because of the specificity of each kind to one type of soil. The commercially available enzymes are listed in Table 9.6. 

---