Lipase chemical modification

Not all ILs are good solvents for proteins, however. There is the interesting example of lipase. Lipase is soluble in both aqueous and organic solvents, so it can be easily solubilized in ILs. Certain lipases even become dispersed or dissolved in some ILs. Since lipase is a very stable enzyme, it catalyzes the hydrolysis of lipids. Enzymatic activity is reported to be maintained in ILs [1]. There is not much published on the solubilization of biomaterials in ILs. In the present chapter we introduce a procedure to use in solubilizing biomaterials in ILs. First we consider the preparation of the IL, and then the chemical modification of biomaterials suitable for dissolution. We have found this procedure helpful when we tried to use electrochemicaUy active biomaterials in ILs. 

Chemical modification of the single free sulfhydryl group of gastric lipases, with either 4,4 -dithiopyridine or 5,5 -dithiobis(2-nitrobenzoic acid) [83, 84], induced a complete loss of activity, when water-soluble p-NPA or emulsified tributy-roylglycerol (tributyrin) were used as substrates [85]. Both catalytic activities were... 

Luthi-Peng, Q., Maerki, H.P. and Hadvary, P. (1992) Identification of the active-site serine in human pancreatic lipase by chemical modification with tetrahydrolipstatin. FEBS Lett. 299, 111-115... 

Chemical Modification of Lipases. The chemical modification of enzymes involving the formation of covalent bonds are a major tool for elucidating the mechanisms of enzymatic catalysis [496 98]. These investigations were aimed primarily at defining those amino acids which participate in catalysis and those which are important in substrate binding. Furthermore, the properties of the enzyme such as solubility, pH optimum, inhibition patterns, and the relative reactivity towards different substrates - the specificity - can be varied by chemical modification. More recently, it was also shown that the enantioselectivity of a lipase may also be improved by covalent modification [499-501] (compare Scheme 2.72 and Table 2.2). 

It is well known that various parameters (e.g. solvent, pH, immobilization, chemical modification and temperature) can have an effect on the enantioselectivity of enzyme-catalysed processes. Most studies in this respect have been carried out on hydrolytic enzymes, especially lipases, esterases and proteases [28]. Recent reports, especially those involving non-hydrolytic enzymes, are discussed below. 

Enhanced activity of an immobilized lipase promoted by site-directed chemical modification with polymers. Process Biochem., 45 (4), 534-541. 

Mesoporous silica thin film served as catalyst support layer for [167] immobilizing lipase without complex chemical modification hydrolysis reaction of 4-nitrophenyl acetate by immobilized lipase followed Michaelis-Menten kinetics... 

B.C. Koops, H.M. Verheij, A.J. Slotboom, M.R. Egmond, Effect of chemical modification on the activity of lipases in organic solvents. Enzyme Microb. Tech., 25 (7) 622-631,1999. 

Two types of interesterification are presently in use chemical and enzymatic. Enzymatic modifications rely on the use of random or regiospecific (1,3-or 2-specific) and fatty acid-specific lipases as catalysts, whereas for chemical modifications metal alkali catalysts are usually employed. 

W.V. Tuomi, R.I. Kazlauskas, Molecular basis for enantioselectivity of lipase from Pseudomonas cepacia toward primary alcohols. Modeling, kinetics, and chemical modification of Tyr29 to... 

DIPE as solvent [168]. The access to both (R)- and (S)-enantiomers was elegantly developed by the enzyme-catalyzed KR. Similarly, the lipase-catalyzed acetylation of 3-hydroxy-3-(2-thienyl)propanenitrile using PSL and later chemical modifications led to the synthesis of duloxetine enantiomers, which are also antidepressant agents [169]. The DKR of the p-hydroxynitrile precursor of duloxetine reported by Backvall and coworkers has allowed the improvement of the overall yield for the synthesis of these drug enantiomers (Figure 9.17) [170]. 

Zeolite membranes are amenable by surface modification with a variety of chemical functional groups using simple silane chemistry, which may provide alternative surface chemistry pathways for enzyme immobilization. In this context, Shukla et al. [238] have recently used a chemically modified zeolite-clay composite membrane for the immobilization of porcine lipase using glutaraldehyde to provide a chemical linkage between the enzyme and the membrane. The effects of pH, temperature, and solvent on the performance of such biphasic zeohte-membrane reactors have been evaluated in the hydrolysis of olive oil to fatty acids. 

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